Molecular Advances to Combat Different Biotic and Abiotic Stresses in Linseed (Linum usitatissimum L.): A Comprehensive Review

Flax, or linseed, is considered a “superfood”, which means that it is a food with diverse health benefits and potentially useful bioactive ingredients. It is a multi-purpose crop that is prized for its seed oil, fibre, nutraceutical, and probiotic qualities. It is suited to various habitats and agro-ecological conditions. Numerous abiotic and biotic stressors that can either have a direct or indirect impact on plant health are experienced by flax plants as a result of changing environmental circumstances. Research on the impact of various stresses and their possible ameliorators is prompted by such expectations. By inducing the loss of specific alleles and using a limited number of selected varieties, modern breeding techniques have decreased the overall genetic variability required for climate-smart agriculture. However, gene banks have well-managed collectionns of landraces, wild linseed accessions, and auxiliary Linum species that serve as an important source of novel alleles. In the past, flax-breeding techniques were prioritised, preserving high yield with other essential traits. Applications of molecular markers in modern breeding have made it easy to identify quantitative trait loci (QTLs) for various agronomic characteristics. The genetic diversity of linseed species and the evaluation of their tolerance to abiotic stresses, including drought, salinity, heavy metal tolerance, and temperature, as well as resistance to biotic stress factors, viz., rust, wilt, powdery mildew, and alternaria blight, despite addressing various morphotypes and the value of linseed as a supplement, are the primary topics of this review.


Introduction
Flaxseed (Linum usitatissimum L.) is one of the oldest cropsand has been farmed for a long time. In Latin, its name translates to "extremely beneficial" crop, presumably because it is a source of three main bioactive components, i.e., α-linolenic acid, lignans, and dietary fibre [1]. This important crop is an annual herb that produces golden-yellow-to reddishbrown-coloured small, flat seeds. In the beginning, the crop was introduced to the US, predominantly to create textile fibres [2]. The stem creates robust, long-lasting, high-quality fibres, which is perhaps one of the reasons that it was named flaxseed. Flax produced from this crop was primarily utilised in the production of textiles (linen) and paper industries. The name linseed is generally used in terms of oil production and other byproducts, like animal feed [3,4].
All varieties of plants responded the most negatively to alkaline-salt stress, and higher concentrations of this treatment completely prevented germination. When five linseed genotypes were developed after being treated with neutral salt and alkaline salt, it was discovered that they had greater salt tolerance [98]. Digital gene expression in flax plants under alkaline-salt stress, neutral-salt stress, and alkaline stress was examined by Yu et al. [99]. In their experiment, it was discovered that under neutral-salt stress, carbohydrate metabolism was impacted, while under alkaline-salt stress, photosynthesis and thereaction to biotic stimuli were badly compromised. In some of the studies, the differential expression of important factors was implicated in responses to abiotic stressors, like WRKY, abscisic acid (ABA), ion channels, and mitogen-activated protein kinase (MAPKK) [10,59]. More of these differentially expressed genes were triggered by alkaline-salt stress than by alkaline stress or neutral-salt stress, indicating that a greater number of genes are involved in regulating the alkaline-salt stress pathway.
Salinity stress has a molecular foundation that is still poorly known. Therefore, Yu et al. [100] used deep sequencing to analyse small RNAs and the degradome in samples that had been exposed to three stress conditions in order to understand the genetic basis of salinity tolerance in flax. Small RNA target genes that control reactions to cues were discovered to be induced. Using transcriptome analysis and degradation genome sequencing, 29 showed opposite expression patterns. The development of climate-smart flax is progressingforall stressors, and 243 miRNA-target combinations have been identified. Two miRNAs, miR398a and miR530, have been reported to be linked to salt stress tolerance in flax. Furthermore, in a separate experiment, Wei et al. [101] discovered that flax seedlings of HIZ019, YOI254, and Tianxin3 had higher salt tolerance because of some important biochemical alterations in flax seedlings under salt stress.

Heavy Metal Stress Tolerance
Heavy metal buildup in vegetation is caused by heavy metal contamination of the earth. Heavy metals in the dietary chain are consequently biomagnified. One such heavy metal that can be detrimental to both plants and animals, even in minute quantities, is cadmium. Plants have evolved specialised regulatory mechanisms that allow them to bind metals to compounds like phytochelatins and metallothionein to create complexes (metallothionein group III). Numerous studies have demonstrated that linen clothing tolerates cadmium pollution well. Even though the toxic effect of cadmium restricts its use for both food and medicine, the use of metal-accumulating plants for the phytoremediation of contaminated soils opens a novel and hopeful path towards improving the resilience of their cultivars and variants to Cd stress [59]. Over the past ten years, extensive research has been conducted on the mechanisms that drivecadmium tolerance in plants. High Cd concentrations reduced root development and, to different degrees, increased the generation of membrane permeability, hydrogen peroxide (H 2 O 2 ), protein oxidation, and lipid peroxides. Additionally, it was discovered that there had been a substantial change in the effectiveness of antioxidant and scavenging enzymes.
The impact of salicylic acid on the antioxidant defence system in flax plants was examined by Belkadhi et al. [102]. Salicylic acid, on the other hand, was discovered to lessen the toxic effects of Cd on the lipid composition of membranes, the antioxidant system, and root development [11]. Belkadhi et al. [103] discovered that a preliminary treatment with salicylic acid preferentially safeguarded plastidial lipids due to greater amounts of polyunsaturated fatty acids in flax. These findings imply that under Cd stress, salicylic acid-pretreated flax plantlets had increased membrane integrity. Salicylic acid boosts root antioxidant mechanisms in flax. Kaplan et al. [104] investigated the effects of cadmium stress on the fatty acid composition of flax in the presence of mycorrhizal fungi. They discovered that seeds from plants cultivated in mycorrhizal fungi contained more unsaturated (18:1, 18:2, and 18:3) fatty acids overall. At 15 ppm of Cd, these impacts became more obvious (the amounts of 18:1, 18:2, and 18:3 were increased by 169,370, and 150%, respectively). These findings indicate that once the level of Cd in seeds hits a certain level, Genes 2023, 14, 1461 6 of 47 this heavy metal increases the effectiveness of the enzymes that control the transformation of saturated fatty acids into unsaturated fatty acids.
Although zinc (Zn) is a crucial ingredient needed for the healthy growth and development of plants, too much of it can also be harmful. Significant attempts have been made to comprehend the tolerance of plants to zinc in flax. Grant et al. [105] discovered that flax cultivars are better at transferring Zn through shoots to seeds than Cd. The use of plant species for phytoremediation is determined by the transfer of metal ions to seeds. Plants are also using a variety of biotechnological techniques to limit the absorption or transfer of the metal ions in plant cells. Smykalova et al. [106] investigated how flax varieties could collect and move Ca and Zn in an in vitro culture made from hypocotyl tissues. The flax varieties discovered by Smykalova et al. [106] are those that are tolerant to zinc, and increasing the amount of zinc and decreasing the amount of cadmium in the kernel will be crucial for increasing the nutritional value of linseed.
The toxic effects of heavy metals like copper (Cu), zinc (Zn), cadmium (cd), nickel (Ni), lead (Pb), chromium (Cr), arsenic (As), and cobalt (Co) on seed germination in various varieties of flax have been assessed in another important research conducted by Soudek et al. [107]. Various heavy metals manifested their harmful effects in the following order: As > Cu > Cd > Co > Cr > Ni > Pb > Zn. In fact, a lot of these studies point to the effective use of flax's potential for phytoremediation, especially when cultivated for fibre production. The growing of flax on heavy-metal-contaminated soils may be made easier with a thorough grasp of the mechanisms underlying heavy metal tolerance [59].

Cold Stress Tolerance
Many cool-season perennial crops' outputs are influenced by two significant physiological processes: vernalisation and photoperiodism. Darapuneni et al. [108] examined the flowering response of various flax genotypes under two photoperiod and vernalisation conditions in growth chamber research. The findings imply that the photoperiod, vernalisation, and genetics significantly influence early blooming in flax. For frigid climate regions like the Upper Midwest of the US and Canada, the early-blooming trait is more important. However, owing to the hot spring and summer temperatures in some places, such as Texas, flax is produced in the autumn. Flax varieties demonstrated genotypic interactions with the photoperiod and vernalisation.
In particular, Texas flax genotypes of the winter variety were susceptible to both vernalisation and blooming photoperiods. The flowering times of most other spring-grown flax genotypes were unchanged by the vernalisation treatments, whereas Texas genotypes postponed anthesis for 7 days or longer in unvernalised seedlings [10]. The majority of other genotypes were unaffected by circadian rhythms (photoperiodism)in vernalised seedlings, while Texas cultivars delayed anthesis for a period of twelve days or longer under vernalised and short-day conditions. Screening for vernalisation and photoperiodic sensitivity in Texas genotypes, as well as the introgression of these characteristics into newly adopted spring-grown genotypes, is required for the development of high-yielding flax genotypes for production sites in the southern Great Plains [59].

Heat Tolerance
Particularly in tropical and subtropical climates, heat stress negatively impacts physiological processes, development (Figure 1), growth, and yield [109]. According to several studies [110][111][112][113], a prolonged duration of heat stress (40 • C for 5-7 days) in relation to flowering duration may have a substantial effect on flax's boll development, pollen viability, pollen production, flowering, seed oil quality, quantity, and seed set. Flax fibre does not need high temperatures. Flax grows best and produces the highest-quality fibre under circumstances that are damp, overcast, and moderately chilly (18-20 • C). High temperatures, especially terminal heat, which is limiting for flax growth, cause poor adaptation of superior fibre linseed genotypes to hotter climates. Although there have not been many

Biotic Stresses
Biotic stresses are responsible for major yield losses [115]; fungal infections, a few viruses, and a phytoplasma are the principal causes of flax biotic stresses. Neither bacteria nor nematodes are known to cause major stress in flax. All varieties of flax (L. usitatissimum L.), such assolin flax (low linolenic acid/vegetable oil flax), linseed (industrial oilseed flax), and fibre flax, are susceptible to fungi. The varieties of each species of flax may react differently to certain diseases or races of the same pathogen. In the parts of the globe where flax is grown, flax diseases vary in frequency, severity, and relevance from one location to another. The most feasible and effective method for engineering disease resistance in crop species is through breeding. Early attempts to enhance flax focused on breeding disease resilience. The morpho-physiological symptoms after biotic stresses in linseed are the yellowing of leaves, a reduced number of seeds, premature ripening (leading to yield loss), and early plant death [116]. Three types of resistance have been found according to the number of genes that influence inheritance: Resistance can be controlled by one gene (monogenic), a few genes (oligogenic), or several genes (polygenic) [117].

Rust Resistance
Flax-growing places around the globe have documented cases of rust. Melampsora lini (Ehrenb.) Desm., a fungal organism, is the causal agent of rust in flax crops. Rusts have the potential to be the most destructiveplant diseases, and flax rust is no exception [118]. Early infections and fast disease development, which are made more likely by meteorological conditions, can totally decimate flax plants and result in a significant reduction in the production and quality of both fibre and seeds. The main difficulty in producing

Biotic Stresses
Biotic stresses are responsible for major yield losses [115]; fungal infections, a few viruses, and a phytoplasma are the principal causes of flax biotic stresses. Neither bacteria nor nematodes are known to cause major stress in flax. All varieties of flax (L. usitatissimum L.), such assolin flax (low linolenic acid/vegetable oil flax), linseed (industrial oilseed flax), and fibre flax, are susceptible to fungi. The varieties of each species of flax may react differently to certain diseases or races of the same pathogen. In the parts of the globe where flax is grown, flax diseases vary in frequency, severity, and relevance from one location to another. The most feasible and effective method for engineering disease resistance in crop species is through breeding. Early attempts to enhance flax focused on breeding disease resilience. The morpho-physiological symptoms after biotic stresses in linseed are the yellowing of leaves, a reduced number of seeds, premature ripening (leading to yield loss), and early plant death [116]. Three types of resistance have been found according to the number of genes that influence inheritance: Resistance can be controlled by one gene (monogenic), a few genes (oligogenic), or several genes (polygenic) [117].

Rust Resistance
Flax-growing places around the globe have documented cases of rust. Melampsora lini (Ehrenb.) Desm., a fungal organism, is the causal agent of rust in flax crops. Rusts have the potential to be the most destructiveplant diseases, and flax rust is no exception [118]. Early infections and fast disease development, which are made more likely by meteorological conditions, can totally decimate flax plants and result in a significant reduction in the production and quality of both fibre and seeds. The main difficulty in producing resistant cultivars is the regular emergence of new races [119] of the pathogen. The availability of genetic resistance sources for most flax-breeding programmes globally has been credited with the disease's rather good global management.
Bright orange and powdery pustules that appear on leaves, stems, bolls, and other aerial plant parts are the primary indications of rust in flax. On leaves and bolls, these pustules are often round, although they are elongated on the stems. Depending on the interplay between a person's ethnicity and genetic makeup, chlorotic or necrotic zones may develop surrounding the pustules. Severe outbreaks result in the leaves drying out and withering, severely defoliating the plants. Numerous urediniospores are produced by rust pustules, and they easily detach and travel long distances in the wind. The black telia pustules that develop from the maturing plants' orange pustules (uredia) create hardy teliospores that can survive the winter [120].
In contrast to many other rusts that need different hosts, flax rust is an autoecious rust, meaning that the fungus may go through all four phases of its life cycle-pycnia, aecia, uredia, and telia-on the flax plant ( Figure 2). Field inspectors seldom ever see the pycnia and aecia, which typically occur on cotyledons and lower leaves early in the season. The stages of pycnia and aecia are crucial for the fungus to complete its sexual life cycle and for the emergence of new races. The uredia are this fungus' third developmental stage. Since uredia develop cycles of urediniospores that might lead to new infections with each cycle during the growing season, the uredial stage is the most harmful to the crop. When the fourth stage reaches maturity, it generates tough overwintering teliospores that can endure inclement weather [120]. resistant cultivars is the regular emergence of new races [119] of the pathogen. The availability of genetic resistance sources for most flax-breeding programmes globally has been credited with the disease s rather good global management. Bright orange and powdery pustules that appear on leaves, stems, bolls, and other aerial plant parts are the primary indications of rust in flax. On leaves and bolls, these pustules are often round, although they are elongated on the stems. Depending on the interplay between a person s ethnicity and genetic makeup, chlorotic or necrotic zones may develop surrounding the pustules. Severe outbreaks result in the leaves drying out and withering, severely defoliating the plants. Numerous urediniospores are produced by rust pustules, and they easily detach and travel long distances in the wind. The black telia pustules that develop from the maturing plants orange pustules (uredia) create hardy teliospores that can survive the winter [120].
In contrast to many other rusts that need different hosts, flax rust is an autoecious rust, meaning that the fungus may go through all four phases of its life cycle-pycnia, aecia, uredia, and telia-on the flax plant ( Figure 2). Field inspectors seldom ever see the pycnia and aecia, which typically occur on cotyledons and lower leaves early in the season. The stages of pycnia and aecia are crucial for the fungus to complete its sexual life cycle and for the emergence of new races. The uredia are this fungus third developmental stage. Since uredia develop cycles of urediniospores that might lead to new infections with each cycle during the growing season, the uredial stage is the most harmful to the crop. When the fourth stage reaches maturity, it generates tough overwintering teliospores that can endure inclement weather [120].

Fusarium Wilt Resistance
A typical flax biotic stress is Fusarium wilt, which is brought on by Fusarium oxysporum. The fungus kills plants shortly after they appear, while later infestations result in the wilting and yellowing of foliage [121]. Agrawal et al. [122] demonstrated that flax wilt resistance is acquired by recessive alleles based on F2 segregation ratios among nine intervarietal crosses comprising the flax wilt-resilient cultivars R552 and RLC6, along with four susceptible genotypes. Spielmeyer et al. [123] investigated the transfer of Fusarium wilt resistance in a recombinant double-haploid (DH) population produced from F2 seeds of a wilt-resistant (twinning Linola TM) and a wilt-prone (Australian flax cultivar Glenelg) line. The partition of two separate genes with additive effects, which accounted for 38 and

Fusarium Wilt Resistance
A typical flax biotic stress is Fusarium wilt, which is brought on by Fusarium oxysporum. The fungus kills plants shortly after they appear, while later infestations result in the wilting and yellowing of foliage [121]. Agrawal et al. [122] demonstrated that flax wilt resistance is acquired by recessive alleles based on F 2 segregation ratios among nine intervarietal crosses comprising the flax wilt-resilient cultivars R552 and RLC6, along with four susceptible genotypes. Spielmeyer et al. [123] investigated the transfer of Fusarium wilt resistance in a recombinant double-haploid (DH) population produced from F 2 seeds of a wiltresistant (twinning Linola TM) and a wilt-prone (Australian flax cultivar Glenelg) line. The partition of two separate genes with additive effects, which accounted for 38 and 26% of the genetic diversity for Fusarium wilt resistance, was considered responsiblefor the phenotypic variance.
Spielmeyer et al. [124] used an amplified fragment length polymorphism (AFLP) marker to assign QTLs to two linkage groups. The flax varieties Mogilevsky-2, Torzhoksky 85, Kievsky, Kalininsky, Rodnik, Nika, K-65 Niva, T-17, and Ustiensky were discovered by Portyankin and Karachan [125] to be resistant to Fusarium wilt. Flax accessions from East Asia and North and South America had greater than average and mean resistance to Fusarium wilt, respectively, but flax accessions from Europe and the Indian subcontinent had lower than average resistance. This variation in resistance to Fusarium wilt was documented by Diederichsen et al. [126].

Alternaria Blight Resistance
Most of the world's flax-growing regions have documented flax disorders linked to infection by Alternaria spp. [127]. Most reports come from India [128] and European countries where the crop is produced. The disease damages leaves and flowers as well as seedlings, producing seedling blight. In India, yield losses of up to 90% have been documented [129], and in the UK, yield losses of up to 35% have been documented [130].
According to Vloutoglou [131], the three most common species on flax are Alternaria infectoria Simmons, A. linicola, and A. alternata. Additional species include A. linicola Skolko and Groves, which is endemic to India and loves temperatures ranging from 26 to 33 degrees Celsius, as well as humidity [132]. In flax waste on and in the soil, A. linicola persists as thick-walled chlamydospores [131]. Flax seedlings are infected by early-season conidia. Infected seeds can spread the fungus, which results in seedling blight [131]. The seedling blight or brown rot caused by the seed-borne inoculum is the first sign of A. linicola. Stunted seedlings that have dark red lesions on their hypocotyls and cotyledons collapse completely in one to two weeks. A rich reservoir of inoculum is created by the diseased and dying seedlings, which spreads to infect healthy plants. On diseased leaves, dark brown lesions develop and typically combine to cover the whole leaf, which then becomes chlorotic and then dies [120].

Powdery Mildew Resistance
Powdery mildew, another significant flax disease caused by the fungus Oidium lini, was first observed in Western Canada in 1997 [133]. Ashry et al. [134] examined flax cultivars for resilience to powdery mildew. They discovered that choosing flax as a fibre variety leads to a rise in powdery mildew incidence. Choosing seed-type flax has the opposite effect, lowering resilience to powdery mildew. When Rashid and Duguid [135] investigated the genetics of resistance to powdery mildew in flax plants, they discovered that resistance to the fungus is caused by a single dominant gene (PM1). The output of flax crops is also known to be impacted by the Pasmo illness. In Western Canada, Septoria linicola (Speg.) Garassini has been reported to cause Pasmo disease and is common and pervasive on flax [136]. Most varieties of flax in Canada are vulnerable, and little is known about how S. linicola isolates are pathogenically dangerous. High-humidity and high-temperature regions show rapid illness progression. S. linicola thrives on agricultural leftovers that have been infected and left in the field from one season to the next.

International
Dillman [46] underlined the importance of collecting germplasm during initial linseed breeding. Further, Zhuchenko and Rozhmina [137] described the use of landraces for fibre flax breeding. Over 48,000 flax accessions are thought to be present in ex situ collections globally, with 10,000 of them possibly being unique. The range of genetic variety maintained in the separate flax national collections (national inventory) of each unique European country, as well as the breeding techniques utilised, are essential for successful breeding activity. The European Cooperative Programme for Crop Genetic Resources (ECP/GR) network adopted the first step towards generating an inventory of the European flax gene pool at an informal meeting of flax-gathering owners within the recently built the Industrial Crops and Potato Network, among the most recent European System of Cooperative Research Networks in Agriculture (ESCORENA), held in Prague on 7 and 8 December 2001 [138].
The 2001 inventory of European flax genetic resources revealed the status. The collection of Bulgarian flax is housed in two institutions. A total 945 accessions were kept at Sadovo in 2001. Of the 450 studied accessions, 23% were linseed, 29% were flax, and 46% were of an intermediate kind. Old and landrace cultivars provided 58% of the total, followed by modern cultivars (15%) and breeder's lines (14%). There were 12 wild species of Linum in the collection other than L. usitatissimum, namely, L. austriacum, L. bienne, L. flavum, L. punctatum, L. grandiflorium, L. perenne, L. setaceum, L. strictum, L. trigynum, L. humile, L. altaicum, and L. viscosum [139]. Moreover, the AgroBioInstitut (ABI) in Kostinbrod maintained 283 accessions. Outof these, 178 came from IgreSadovo, 29 were from other Bulgarian institutes, 71 were from abroad, and 5 were self-bred specimens. These accessions came from Europe, Asia, Africa, and America, respectively. The collection is made up of 54% cultivars, 27% landrace and primitive cultivars, 15% breeding lines, 1% wild forms, and 2.5% genetic stocks. By plant type, the collection was constituted by 31% fibre flax, 35% linseed, 32% combination type, and 2% other types. According to the International Union for the Protection of New Varieties of Plants [140,141], the International Flax Database (IFDB) descriptors [142,143], and other species descriptors, 259 accessions for 15 morphological traits, 5 biological characteristics, and 4 yield traits were described [144,145].
The assessment, description, and characterisation systems for flax germplasm vary depending on the needs of breeders in each country, whether it is in Europe, the USA, or Canada. To exploit the genetic diversity of flax for breeding purposes, a good, unique approach for evaluation, description, and characterisation is necessary, which should be standard at least across European nations. This is because different evaluation and characterisation techniques are employed in various European genebanks as well as outside. Thus, the IFDB may be viewed as the first input for this goal [143]. The IFDB was initially overseen and managed by Agritec Ltd. (Viale delle Terme di Caracalla, 00153 Roma (Italy)) starting in1993 in the framework of FAO ESCORENA Flax and Other Base Plants Network (FAO FOBPN) [146] and then, over time, the framework of IPGRI Coordination Group Network for Sugar, Starch and Fibre Crops (CGNSSFC), currently namedSugar, Starch, Fibre Crops & Aromatic Plants Network (CGN SSFC&APN) at Bioversity International. In theACCESS IFDB framework, it currently contains passport information for 8385 accessions of 11 collections from 10 nations. It is estimated that 37% of the accessions are unique based on a study of the provided data. It also showed that there were significant variations in the collections' description fill rates. Table 1 displays the findings of current germplasm characterisation and assessment research carried out at the Plant Gene Resources of Canada (PGRC), Canadian National Seed Gene Bank [147].

National
Cultivated forms have been reported to be rich in diversity in the Indian region. A total of 2627 accessions, comprising 2495 indigenous, 51 released, and 132 foreign varieties, were preserved in long-term and medium-term storage at the National Bureau of Plant Genetic Resources (NBPGR), New Delhi, India. According to the information shown below, the Central Variety Release Committee (CVRC) and States Variety Release Committee have released and notified 51 improved varieties for general cultivation in various agro-climatic conditions. Table 2 lists the different varieties of linseed grown in different climatic regimes of India [147]. Table 3 contains list of linseed varieties that are tolerant to various abiotic and biotic stresses. Cultivars with dual purpose (seed + fibre) Him Alsi-2, Nagarkot, Jeevan, RLU-6, Rashmi, Parvati, Meera, Shikha, Tiara (JRF 2) Table 3. List of linseed varieties that are tolerant to various abiotic and biotic stresses.

S.
No.

Name of Variety
Year of Release

Breeding through Conventional Methods
Collections of flax and linseed genetic resources are researched for breeding purposes, and the selection of the first practical parental patterns is commonly debated. According to Tadesse et al. [148], who conducted research on crossings between high-yielding and lowyielding flax components, it may be beneficial to hybridise low-and high-yielding flax lines in order to increase genetic diversity and produce high-yielding flax lines. According to the findings of Pavelek [149], crosses of high-yielding flax components may be productive when the genetic variability of the parental lines is substantial. The largest linseed growers and producers in the world are India, the USA, and Canada. Consequently, increasing seed production, increasing oil content and quality, snowballing resilience to diseases and pests, and developing early-maturing accessions are the key breeding objectives.
Today's breeding efforts are mostly focused on traits that stabilise yields, such as resistance to fungal infections, and qualitatively novel contents of materials. Mutagenesis [150,151], genomic approaches [152], and other methodsare used to develop new breeding techniques; for example, anther culture, mutation breeding, molecular-markerbased analysis [153], and techniques of transformation can be employed to find a solution when the genetic variability's options have been exhausted and are no longer available. For breeding fibre flax, the most crucial factors are fibre content and resistance to Fusarium and scorch, whereas for linseed, the most crucial factors are seed production, fatty acid composition, and resistance to Fusarium and rust [154].

Interspecific Hybridisation
Although many interspecific crossings have been made, only those involving species with a similar number of chromosomes have so far proved beneficial. The F 1 offspring of hybrids between L. usitatissimum and a few other species with n = 15 chromosomes are robust and fruitful, and they have been exploited to introduce certain desired traits, such as resistance to linseed rust.
Through modifications to the pollination of the cultivated species, interspecific hybridisation can also increase genetic variability. Some Linum species, including L. grandiflorum, L. anstriacum, and L. perenne, are incompatible with one another. Numerous attempts were made to cross the cultivated linseed (L. usitatissimum) with the self-incompatible species without success. When L. usitatissimum was employed as the female parent in crosses, abortive seeds that were dried up and unable to germinate were obtained [155]. The embryo of the cross between L. usitatissimum and L. grandiflorum degenerates after 7 days due to somatoplast sterility, according to embryological investigations. By cultivating the embryos of the cross and transmitting the incompatibility mechanism to the cultured species, attempts are being made to raise the hybrid. Induced self-incompatibility would force cross-pollination and raise genetic variation in farmed plants [147].

Hybrid Linseed
Knowledge of floral morphology and pollination methods is necessary to produce hybrid varieties with vigour or heterosis effects. Flax heterosis has ranged from 40% [156] to 231% [157]. The development of flax and linseed hybrids is less common than that of sorghum, maize, sugar beet, onions, or pear millet. The primary cause is that the male sterile flax flower has a tiny corolla that does not open to facilitate cross-pollination. Several researchers have investigated the issue of the male sterile flax, including Kumar and Singh [158,159], Dubey and Singh [160,161], and Kumar [162]. Particularly in India, several researchers have looked at the compatibility of flax or linseed kinds in diallel crosses, allowing for the assessment of the heterosis impact. Pant and Mishra [163] found positive and substantial heterosis for seed yield per plant at 40 crossings and fibre yield at 13 crosses. This impact frequently results from dominance, non-allelic interaction, or the combined influence of additive genes.
Trait selection is influenced by the relationship between the genes expressed by either the general combining ability (GCA) or the specialised combining ability (SCA). In contrast to the nonadditive effects of SCA, which were confirmed by Sood et al. [164] for number of seeds per capsule and seed weight, the GCA of flax and linseed is related to traits like the number of capsules per plant, plant height, the weight of capsules per plant, and seed yield. There were occasionally both additive and nonadditive gene effects seen [165].

Mutation Breeding
Despite Tammes reporting a mutation in linseed as early as 1925, nothing is known about the nature, induction, and use of induced mutations in flax. Mutagenesis in linseed has been attempted using both physical [166,167] and chemical mutagens [168,169]. Deshpande [170] identified flax seedlings with low levels of chlorophyll in the NP-12 normal group. Levan [171] discovered three families segregating for chlorophyll mutations in diploidplants but not in tetraploid plants following X-irradiation. Three different linseed varieties, Hira, Mukta, and Neelum, were exposed to radiation of10, 20, 30, and 40 kR by Rai and Das [167].
Along with various kinds of chlorophyll mutations, an increase in mutation frequency was seen with increasing irradiation dosages. The observedcholorophyll mutants showed delayed blooming, shorter plants, and fewer capsules per plant. In contrast to normal plants, these mutants had a much higher number of non-bearing tillers per plant. Sharov [172] reported mutants with good yield and a 52-56% increase in resistance to F. oxysporum following chemical mutagenic therapy. After irradiating the variety Neelum, George and Nayar [166] acquired a dwarf mutant (TL-1), which developed 30 days quicker than the typical plant. It is interesting to note that the mutants had larger 1000-seed weights and lighter-coloured oils with higher oil contents.
In Australia and New Zealand, three new linseed cultivars, known as "Wallaga", "Argyle", and "Eyre", have been introduced under the generic name "Linola". Goldenyellow seeds are seen in "Linola" varieties. "Eyre" is a result of a single-plant selection made in the F 4 of the cross "Glenelg"/CPI 84495//4*"Zero", from which an F 8 bulk was obtained. The EMS (ethyl methane sulphonate) mutagenesis of the Australian linseed cultivar "Glenelg" and the recombination of two mutant genes produced the low-linolenic acid genotype "Zero" [173].
Eight Indian states, namely, Madhya Pradesh, Bihar, Gujarat, Delhi, Punjab, Rajasthan, Uttar Pradesh, and West Bengal, are focused on linseed breeding [174]. India's linseed breeding industry is well established and well organised. The Indian Central Oilseeds Committee (ICOC) increased its linseed research activities in 1947, and then in 1958, the project "Intensification of Regional Research on Cotton, Oilseeds and Millets" (PIRCOM) took over. The All India Coordinated Research Project on Oilseeds (AICRPO) was launched by the Indian Council of Agricultural Research in 1968 [174] utilising international methods.

Molecular Diversity
Flax genetic diversity can be assessed using a variety of methods, including physical traits, isozymes, and molecular markers. While isozymes are scarce, morphological characteristics are frequently more quantitative and reliant on their surroundings. The prevalence and distinctiveness of DNA markers are some of their benefits. They are helpful for identifying varieties and assessing DNA diversity. To examine the genetic variation in flax, various molecular markers, such as restriction fragment length polymorphism (RFLP), random amplified polymorphic DNA (RAPD), amplified fragment length polymorphism (AFLP), and simple sequence repeat (SSR), have been developed.
By using RAPD markers, Fu [29] evaluated the geographic trends of flax variability in the global collection of farmed flax. A total of 149 RAPD markers were measured for each of the 2727 flax accessions, which came from 63 different nations and 1 unidentified group. The accessions were divided into 12 main areas, which explained 8.2% of the RAPD variation. The most varied accessions came from East Asia and European areas, but the most distinctive accessions came from the Indian peninsula and Africa. Genetically, accessions from the West Asia area were less related to those from the Indian peninsula and more related to those from Africa. These discoveries are crucial for comprehending the domestication of flax and are also helpful for categorising the intraspecific variety of cultivated flax, identifying the fundamental subgroup of the flax collection and looking into novel gene sources for flax improvement.
Cloutier et al. [175] conducted the first comprehensive study on the production and analysis of a considerable number of SSR markers in flax. The genetic relationship among 635 genotypes was discovered by using 275 variable EST-SSRs. The pedigree connection between the accessions was associated with subclusters within the major clusters. The expressed sequence tags-signature sequence tags (EST-SSTs) created here signal the beginning of SSR markers in flax on a wide scale. They could be applied to the creation of physical and genetic maps, studies of genetic variation, association mapping, and cultivar identification.
Several molecular methods are presently available to identify germplasm at the DNA level. Due to the large number of markers that can be produced per analysis, AFLPs are typically regarded as being comparatively strong in germplasm analysis when compared to alternative genetic marker systems. In the research by Van Treuren et al. [35], duplicate germplasm in the flax collection of the Netherlands' Center for Genetic Resources was identified using AFLPs. ANOVA was used to compare accessions in pairs in order to find duplicate genotypes. The 29 accessions of breeder's lines could be reduced to 14 by stepwise bulking accessions until all surviving accessions were noticeably distinct. This bulking strategy only had a negligibly tiny negative impact on the among-population component of variance, demonstrating a 2.6% decrease. This outcome is addressed in connection to enhancing collection management effectiveness.

Association Mapping
Association mapping studies are crucial throughout animal and plant research because they aid in the discovery of genes that influence phenotypic variation and create the framework for the recognition of genes that play a vital role in what is seen as variation. Genome-wide association studies (GWASs) have become a more common practice in the quickly developing genomic age. The early accessibility of complete genome sequencing data in humans appears to have contributed to the broad application of GWASs throughout human research before it was adopted by plant science. Several plant genomes have been mapped as a result of the significant advances in sequencing technology over the past ten years. The availability of plant whole-genome sequences gave researchers the chance to perform effective GWASs [59].
Traditionally, biparental crosses, which are very simple to create, have been used to identify quantitative trait loci (QTLs) in plants. Biparental hybrids do have some restrictions. Since the community is descended from two parents, it only works with genetic variance within the parental lines, which is the most important restriction. Additionally, biparental QTL mapping has poor precision. A GWAS, in comparison, has several advantages over QTL mapping, including high resolution and the use of genetic variation present in readily available germplasm resources. The genetic makeup of the targeted material, phenotypic diversity, and the quantity and distribution of molecular markers have a significant impact on the effectiveness of GWAS.
Single-nucleotide polymorphism (SNP) markers have all the characteristics needed for high-throughput genotyping and are widely available and evenly dispersed throughout the complete genome. The majority of GWASson plants are conducted on important species like rice, maize, and wheat, as well as model species like Arabidopsis. Limited attempts have been made to perform GWASs on flax compared to the main food products and model species. Since flax is an oilseed grain, its breeding programme focuses mainly on increasing the seed oil content and total output. Secured output is heavily reliant on climatic factors that exert various biotic and abiotic pressures [59].
Plant resistance against stresses is governed by a sophisticated polygenetic system. GWAS is a cost-effective and effective way to analyse such intricate genomic control. For the recognition of significantly associated loci and subsequent candidate gene identification, an in-depth knowledge of the level of linkage disequilibrium (LD), the profile of LD variations across the entire genome at substantially associated loci, and functional annotations of predicted gene models is required [59].

Extent of Linkage Disequilibrium
The degree of LD differs significantly between various chromosomes and loci on the same chromosome, and it is not consistent across plant species. Population drift, selection, admixture, inbreeding, and taming are some of the factors that influence LD. The rate of recombination, transversion, translocation, and chromosomal duplications all play a significant role in the LD variation at various genetic locations. Numerous plant taxa have been the subject of in-depth research on LD decays. Overall, species have been found to cross-pollinate more frequently and experience very little LD degradation. For instance, it has been found that LD decays occur within 0.3-2 kb [176,177]. However, self-pollinated species like soybean have shown evidence of LD decay after extremely lengthy distances [178]. According to Sonah et al. [178], distinct LD decays at r 2 = 0.2 were observed in soybean across various chromosomes, spanning from 250 kb to 2.5 Mb on average. Similarly, the flax genome exhibits comparatively delayed LD decays relative to cross-pollinated species since it is a self-pollinated plant species. Soto-Cerda et al. [179] used genetic information from 448 microsatellite markers and 407 flax accessions from around the world in a genome-wide analysis. According to Soto-Cerda et al. [180], LD declined across the entire genome within an average of 1.5 cM. Like the previous studies, [181] carried out two additional genome-wide investigations using 460 SSRs for 390 accessions and 112 SSRs for 407 accessions, respectively, and found LD decay up to 1.8 cM. Aside from the availability of the whole-genome sequence, insufficient attempts have been made to carry out an effective LD analysis and consequent GWAS.

Genetic Loci Identified by GWAS
One of the main goals of flax breeding programmes is to increase the amount of linolenic acid (LIN) in the plant. It is difficult for breeders to increase LIN content without influencing other fatty acids (FAs) because LIN content has negative correlations with linoleic acid (LIO), stearic acid (STE), palmitic acid (PAL), and oleic acid (OLE) [182]. Some flax accessions with high LIN were discovered through careful evaluation of a wide range of germplasm, but they are not well suited to cropping systems [183,184]. Identification of the genetic regions controlling FA composition is crucial in this respect. Nine loci that are strongly linked to five seed quality characteristics have been found in flax. For the iodine index, LG8 linkage group (LG), LG5 linkage group (LIN), total oil content, and LG9 linkage group (LG), the loci with the largest impacts have been described. The markers on LG2 (Lu2046) and LG6 (Lu2555), however, described about 8 and 4% of the phenotypic variance in PAL and OLE, respectively. The research was unable to find substantially linked loci for PAL and OLE. The reason that there is a negative association between LIN and other FAs may be explained by several markers associated with various FAs that were discovered to be colocalised in this research. It is interesting to note that Soto-Cerda et al. [181] found a locus linked to genes expected to be involved in FA biosynthesis.
For instance, it was discovered that the acyl-CoA: diacylglycerol acyltransferase A (dgatA) gene is located near the strongly linked locus on LG3. Earlier, a biparental quantitative trait locus (QTL) mapping strategy was used to identify flax dgatA. As a result, the loci found in flax using the GWAS method appear promising.

Molecular Mapping of QTLs
Genetic mapping has been used widely in both animal and plant species since the finding of DNA-based identifiers. After Botstein et al. [185] released the initial report of restriction fragment length polymorphism (RFLP), genetic mapping efforts increased, but they were hampered by the lack of an effective study. Early in the 1990s, advances in mapping analysis led to the first genetic map spanning the complete set of chromosomes, becoming available and being reported in humans and some plant species. The first RFLPmarker-based linkage map was created in 1988 and used for QTL mapping in plants [186]. The first inheritance studies and genetic mapping of flax were conducted in 1930 by Henry, who looked at how rust protection was passed down in the Bombay and Ottawa 770B varieties. These mapping initiatives, however, were based on physical traits. The first genetic linkage map of flax was created using RFLP and RAPD markers following the development of DNA-based molecular marker methods [187]. The 1000 cM genetic linkage map created was made up of 15 unique linkage groups with ninety-four evenly spaced markers. The F 2 mapping population created from the hybrid between CI1303 and Stormont Cirrus was used in the research. Multiple investigations have since discovered QTLs for various flax characteristics. Cloutier et al. [175] constructed a genetic linkage map utilising114 SSR markers and 5 SNPs in a DH population of 78 individuals obtained from a crossing of SP2047 (yellow-seeded withlittle linolenic acid) and UGG5-5 (brown-seeded withelevated linolenic acid). For LIO, LIN, and the iodine value (IOD), two significant QTLs were found for each, and one major QTL was found for palmitic acid. The QTL QPal.crc-LG9 explained 42% of the trait variance in palmitic acid.
In related research, Kumar et al. [188] used a recombinant inbred line (RIL) population of 243 individuals from a hybrid between cultivars CDC Bethune and Macbeth to create a genetic map using 329 SNPs and 362 SSRs. For a total of 14 distinct characteristics, 20 QTLs were found in their study. There were three QTLs for STE and OLE, two QTLs for iodine content and LIO, and two QTLs for seed protein, oil content, thousand-seed weight, cell wall, straw weight, output, seeds per boll, and days to maturity. Candidate genes for characteristics related to yield component traits, cell wall synthesis, fibre production, and fatty acid biosynthesis were discovered through the analysis of the QTL.
Omics-aided technologies have been utilised to conduct stress tolerance research on rice [192], soybean [193], and flax [194]. In contrast to other crops, however, comparatively fewer attempts have been made to use the available genetic and genomic tools for flax development. In accordance with Akhmetshina et al. [195] and Shivaraj et al. [59], who investigated the application of omics-assisted breeding and high-throughput sequencing technologies for the advancement of climate-smart flax, advanced tools such as genomewide association studies (GWASs) and genomic selection in conjunction with various omic technologies offer an opportunity to increase the precision of flax improvement and plant selection.
In order to introduce weather-resilient characteristics into flax cultivars for long-term productivity, an extensive strategy incorporating a variety of technologies can significantly simplify the process [196]. . An overview of the use of omics approach, which depicts how the information is passed on from genomics to transcriptomics, then to proteomics, and finally to metabolomics, which bring about the final change in response to a stress or signal.

Genomics
The genetic diversity of flax was evaluated in the early years of the twenty-first century using a variety of molecular markers [187,[197][198][199][200]. The release of the flax wholegenome transcriptome provided a significant advantage in the creation of genomic tools [201]. The information from reduced representation sequencing and whole-genome re- Figure 3. An overview of the use of omics approach, which depicts how the information is passed on from genomics to transcriptomics, then to proteomics, and finally to metabolomics, which bring about the final change in response to a stress or signal.
Omics-aided technologies have been utilised to conduct stress tolerance research on rice [192], soybean [193], and flax [194]. In contrast to other crops, however, comparatively fewer attempts have been made to use the available genetic and genomic tools for flax development. In accordance with Akhmetshina et al. [195] and Shivaraj et al. [59], who investigated the application of omics-assisted breeding and high-throughput sequencing technologies for the advancement of climate-smart flax, advanced tools such as genomewide association studies (GWASs) and genomic selection in conjunction with various omic technologies offer an opportunity to increase the precision of flax improvement and plant selection.
In order to introduce weather-resilient characteristics into flax cultivars for long-term productivity, an extensive strategy incorporating a variety of technologies can significantly simplify the process [196].

Genomics
The genetic diversity of flax was evaluated in the early years of the twenty-first century using a variety of molecular markers [187,[197][198][199][200]. The release of the flax whole-genome transcriptome provided a significant advantage in the creation of genomic tools [201]. The information from reduced representation sequencing and whole-genome re-sequencing has since been successfully applied to comprehend crop diversification, construct linkage maps, recognise markers, and discover QTLs in flax. Using the genotyping-by-sequencing (GBS) method, 258,873 SNPs spread across fifteen linseed chromosomes have been reported [188].
Recent studies have used genome-wide association studies (GWASs) to find potential SNPs for various traits, including improved stress tolerance indices, yield, and other traits in different species, including sorghum [202][203][204], rice [205], and sesame [206,207]. Although the AQP gene family improves drought resilience, multidrug and toxic compound extrusion (MATE) mediates the reaction to abiotic stresses [208]. However, research success in flax has beenless rapid than in cereals [209,210]. He et al. [211] used GBS to find 258,873 SNPs from the Canadian flax accession (core collection) as part of a GWAS to discover genetic areas linked to Pasmo resistance. Of the 500 potential QTLs found, 45 covered 85 resistance genes. Additionally, two potential genes (Lus10031043 and Lus10020016) for flax protection against this pathogen were found to be similar based on orthology with Arabidiopsis thaliana genes. In recent years, You et al. [212] examined 447 flax accessions for resistance to powdery mildew over 5-8 years from three sites, encompassing 372 samples from the primary collection and 75 mating lines.
By estimating the genetic potential rather than looking for specific QTLs, genomic selection (GS) is a breeding technique that overcomes the restrictions of marker-aided selection (MAS) for pace breeding (Figure 4). GS has the capacity to resolve all variations in the genetics of complex traits, as opposed to the traditional agricultural breeding technique, which is slow in concentrating on the complicated and barely heritable quantitative variables. Because of this, it is becoming clear that using molecular genetic markers to create brand-new models based on markers for genetic assessment is a promising method [196]. The precise phenotyping of a selected diverse collection of genotypes (for the purpose of breeding) in various conditions is required in order to develop a statistical model (GS model) that can be used to calculate genomic-predicted breeding values in the breeding population. As it addresses the minor effects of QTLs, the GS technique has many benefits over traditional and marker-assisted breeding [213].
Because it combines all known molecular markers with phenotypic information, GS has been promoted as the most effective technique for determining genetic values for selection [214,215]. An enhanced genetic gain/unit period was observed during the breeding cycle in flax GS experiments [216,217]. Tables 4 and 5  You et al. [212] used 447 flax inclusions as an experimental population to create the most current model for forecasting the evaluations for powdery mildew across five years at three separate sites. The 349 QTNs discovered through GWAS described ninety-six percent of the variance in powdery mildew, demonstrating the model's strong predictive power and promise for use in genomic prediction.
By estimating the genetic potential rather than looking for specific QTLs, genomic selection (GS) is a breeding technique that overcomes the restrictions of marker-aided selection (MAS) for pace breeding (Figure 4). GS has the capacity to resolve all variations in the genetics of complex traits, as opposed to the traditional agricultural breeding technique, which is slow in concentrating on the complicated and barely heritable quantitative variables. Because of this, it is becoming clear that using molecular genetic markers to create brand-new models based on markers for genetic assessment is a promising method [196]. The precise phenotyping of a selected diverse collection of genotypes (for the purpose of breeding) in various conditions is required in order to develop a statistical model (GS model) that can be used to calculate genomic-predicted breeding values in the breeding population. As it addresses the minor effects of QTLs, the GS technique has many benefits over traditional and marker-assisted breeding [213]. Because it combines all known molecular markers with phenotypic information, GS has been promoted as the most effective technique for determining genetic values for selection [214,215]. An enhanced genetic gain/unit period was observed during the breeding cycle in flax GS experiments [216,217]. Tables 4 and 5 Table 4. Genome-wide association studies discovered quantitative trait nucleotides/loci for key abiotic stressors [196].   Table 5. Genome-wide association studies that discovered quantitative trait nucleotides/loci for key biotic stressors [196].

Lus10031058
Elongation Factor Effector-trigger immunity [135]  -Pm1 [135] Lu4-12,432,479 Lus10036891 RGA (WRKY transcription factor) [212] More comprehensive GS studies are anticipated soon due to the enhanced genetic techniques with genomic resources for linseed, which could help in the release of new cultivars suited to requirements. Due to greater costs than MAS, the more widespread use of GS is currently still a problem. Budhlakoti et al. [216] reviewed the present state of genomic selection research in crop plants and the prospects for its effective application in the making of weather-resilient crops. They stressed that research on genetic makeup under heat with drought stress can invariably speed up the production of stress-resilient cultivars through genomic selection [196].

Transcriptomics
The discovery of important genes engaged in the stress tolerance process is aided by transcriptome profiling, which offers a thorough overview of gene expression and regulation. Depending on the abundance of generated genomic materials and plant variety, different methods are used to investigate the transcriptome, including spotted microarrays, expressed sequence tags, Affymetrix Gene Chips, sequencing combined with suppression subtractive hybridisation, and RNA sequencing. The advancement of next-generation sequencing technologies and RNA sequencing has made them the most efficient, affordable, and high-throughput transcriptomic techniques [196].
Numerous transcriptomic studies have been conducted to date to determine the impact of salt and drought on oilseed products like sesame [207], flax [220], jatropha [221], sunflower [222], and soybean [223]. The sequencing of the linseed genome and the presence of genetic maps [182,195,201] set the groundwork for numerous transcriptomic studies and the discovery of genes underpinning characteristics of economic and agronomic significance. High-throughput sequencing was used to examine how flax responded to the alterations in the levels of salt and pH [224][225][226], drought [227], nutrient deficiency [228,229], and metal stress [230].
Dash et al. [227] performed a transcriptome analysis on the relatively drought-tolerant flax cultivar of India, i.e., T-397, and expression profiling assisted in identifying markers for the identification of drought-resistant flax varieties. Shivaraj et al. [231] showed the increased expression of integral membrane proteins, primarily aquaporins, using transcriptome analysis data, which improved the knowledge of their physiological function. In a transgenic strain of flax cv. Blanka, the overexpression of the DREB2A gene confers drought resistance [73]. Like this, only a small number of flax genes have been identified and functionally characterised under extreme temperature stress. According to Saha et al. [111], 34 potential HSF genes from the flax genome were identified across the entire genome.
Under conditions of heat stress, different expression patterns are bestowed by HSF, along with NAC domain TFs. By lengthening roots, reducing membrane injury, and improving ion transport, two salt-tolerant genes that are related to Arabidopsis Senescence-Associated Gene 29 (SAG29) may improve salt tolerance. The flax transcriptome reaction to acidic soil revealed genes with altered expression patterns [232]. Through transcriptomics, the reaction of flax to an unfavourable zinc shortage and soil acidity identified genes incorporated in cell wall biogenesis photosynthesis and ion transfer [224]. Several pathogen-related dominant genes were induced to counteract the negative impacts of low Zn concentration. Melnikova et al. [228] found 96 conservative microRNA homologs from 21 families, and they also described the function of 7 microRNAs (miR408, miR395, miR168, miR398, miR169, miR399, and lus-miR-N1) in the control of metabolism and gene expression in plants during nutritional stress.
As a defence strategy, plants' transcriptional and translational profiles are also altered by pathogen assault, which causes the implementation of various metabolic pathways and genes. In their analysis of the gene reaction during the early phases of Fusarium infection, Kostyn et al. [233] found 47 genes responsible for antioxidant biosynthesis and phenylpropanoid pathways in flax. Hundreds of genes with differential expression in reaction to early pathogenesis were found in the prevalent Canadian cv. CDC Bethune transcriptome of flax immune to Fusarium wilt and a susceptible variety, Lutea. Several important genes, including those involved in secondary metabolism, the stimulation of pathogenesis-related (PR) interactions, and the formation of lignin, have higher transcript abundance among these [196]. The transcriptome of two susceptible and two resistant BC 2 F 5 populations, as well as four cultivars of fibrous flax, revealed the enhanced expression of many genes involved in the defence response, such as PR protein-encoding genes, ROS production genes, and genes related to cell wall formation [234].
Boba et al. [235] recently found that upon Fusarium infection in flax, upregulation of the terpenoid pathway, resulting in higher ABA content, initiates the fast plant response, and PR genes, particularly chitinase and 1,3-glucanase, play an important role in resistance. In previous research, Wróbel-Kwiatkowska et al. [236] found that transgenic flax plants overexpressing the 1,3-glucanase gene displayed reduced vulnerability to this pathogen. In reaction to infection by Fusarium oxysporumlini, it was discovered that the resistant flax cultivar's transcriptomic response was faster and more effective, enabling the translation ofa greater number of activated and repressed genes [237].
The NCBI Sequence Read Archive and NCBI Gene Expression Omnibus libraries contain the transcriptome sequences, and gene expression data for flax. L. usitatissimum microRNA sequences are provided in the "miRbase" database, along with their main and secondary structures and locations within the flax genome. In the field of flax transcriptomics, there are more studies published (Table 6) on tolerance to abiotic stressors than on resistance to biotic stresses, which is significant and might be a result of targeted characteristics under breeding programmes for areas. Most of the transcriptomic research only looked at one or two genotypes; however, for comparative analysis and gene annotation, a larger number of varied genotypes should be examined [196]. Table 6. Global transcriptome study of gene expression patterns in flax in response to significant abiotic and biotic stressors [196].

Metabolomics
Interpreting the mechanisms of plant biology requires integrating metabolomics because metabolites, which are the consequences of gene expression, provide a detailed picture of an organism's biological and physiological condition [243][244][245]. However, as discussed by Hall et al. [246], metabolomic research is not without its difficulties because the extent of the plant metabolome is unclear. Plants have a widervariety of metabolites than other species. A wide range of statistical channels, involving mass spectrometry (MS)-and nuclear magnetic resonance (NMR)-based techniques like CE-MS (capillary electrophoresismass spectrometry), GC-MS (Gas Chromatography-Mass Spectrometry), FTIR, and LC-MS (Liquid Chromatography-Mass Spectrometry), have been used to identify and quantify awide variety of primary and secondary metabolites produced in response to abiotic and biotic stresses. NMR is usually used to identify moderate-to-high-abundance metabolites, and it requires little sample preparation.
Additionally, new improvements in NMR superconducting magnet field intensity have led to better spectral precision and detection sensitivity. In contrast to NMR, modern MS-based methods produce analyses of complicated plant metabolite mixtures with greater sensitivity. A review paper by Ibáez et al. [247] provides a summary of the most contemporary literature on direct ionisation methods used in the metabolomics of food. It is well known that metabolites like lignans, particularly omega-3 fatty acids and polyunsaturated fatty acids (PUFA), are likely to induce a positive impact on nourishment and the avoidance of some diseases [248,249]. In order to choose flaxseed types with a superior nutrient composition, a tool based on NMR metabolomics was developed [250]. Lignans and omega-3 fatty acids, two bioactive substances of importance for human health, are significant components of flaxseed. Oilseeds have been found to accumulate a wide range of secondary metabolites, such as sugars, inorganic ions, glycine betaine, and proline, in addition to the metabolomics of nutritive compounds, to promote plant adaptation to abiotic stress [251].
BABA, for example, has been shown to play a part in promoting drought resistance in a variety of plants, including Arabidopsis [252], apple [253], rice [254], spring wheat [255], potato [256], and tomato [257]. More osmoprotectants, anthocyanins, and proline were accumulated when PR1, PR2, and PR5 genes were overexpressed in Arabidopsis, while trehalose production made tobacco more drought-tolerant [258,259]. In reaction to water stress, BABA in flax increases proline and nonstructural carbs while decreasing aspartate concentration and inorganic solutes [260]. Flax [261], along with rice, was shown to have relatively higher concentrations of proline and glycinebetaine under salinity stress [262].
Total soluble sugars, compatible solutes, and total protein contents, like proline and betaine, were found to increase with increasing salinity in flax genotypes under PEGinduced water stress and saline-alkaline environments, suggesting that they may play a role in adjusting to osmotic stress [263]. Under salt and/or osmotic stress, different levels of both wild-type and PLR-RNAi transgenic flax, lipid peroxidation, and a distinct metabolic profile of MDA have been noted [261,264]. As a defence strategy, pathogen assault also causes changes in several secondary metabolites, including lignin, flavonoids, polyamines, catecholamines, terpenoids, tannins, phenyl propanoic acids, and phenolics, which are produced as a result of the translational profile of plants. Many rice studies related to metabolomics have been conducted to identify important metabolites and routes in reaction to different types of environmental distress [265]. The purpose of these studies was to comprehend how the model crop rice induced defence mechanisms involving effector-prompted immunity (ETI) and PAMP-triggered immunity (PTI) in response to pathogens. As a result, stress causes significant alterations in a plant's metabolic makeup ( Figure 5), and full metabolite profiling may offer important insights into processes for stress tolerance [196].

Proteomics
Proteomics is the real-time analysis of the molecular and functional properties of every protein in a living thing. It combines different bioinformatic techniques with twodimensional (2-D) gel electrophoresis, Western blotting, mass spectrometry (MS), matrixassisted laser desorption ionisation-time of flight (MALDI TOF), and ELISA [266,267]. Recent advances in proteomics have decreased protein evaluation mistakes and opened new avenues for high-throughput proteome studies. Most of the proteomic research has been undertaken on the entire genomes of soybean, wheat, barley, rice, and potatoes. Reports on proteomic research on oilseeds include those on flax [268,269], sunflower [270], and Indian mustard [271].
Proteome analyses have shown that tolerant plants have consistently increased levels of stress-reactive chains of amino acids compared totheir sensitive counterparts, which include transcriptional regulators like Myb protein, B-Peru-like protein, and SWIB/MDM2 protein involved in anthocyanin biosynthesis [272,273]. In comparison to their sensitive counterparts, drought-tolerant wheat and barley types were found to have higher levels of proteins, such as several chaperons (HSP70, HSP90, and cyclophilin A), glutathione-Stransferase (GST), and lipoxygenase (LOX) [274]. Another studyfound that drought [275], low temperature, and salt [276] caused reductions in RubisCo, enzymes like phosphoglycerokinase (PGK), and transketolase phosphoribulokinase (PRK) in wheat [277]. An increase in numerous minor heat shock proteins, as well as HSP82 from the HSP90 fam-

Proteomics
Proteomics is the real-time analysis of the molecular and functional properties of every protein in a living thing. It combines different bioinformatic techniques with twodimensional (2-D) gel electrophoresis, Western blotting, mass spectrometry (MS), matrixassisted laser desorption ionisation-time of flight (MALDI TOF), and ELISA [266,267]. Recent advances in proteomics have decreased protein evaluation mistakes and opened new avenues for high-throughput proteome studies. Most of the proteomic research has been undertaken on the entire genomes of soybean, wheat, barley, rice, and potatoes. Reports on proteomic research on oilseeds include those on flax [268,269], sunflower [270], and Indian mustard [271].
Proteome analyses have shown that tolerant plants have consistently increased levels of stress-reactive chains of amino acids compared to their sensitive counterparts, which include transcriptional regulators like Myb protein, B-Peru-like protein, and SWIB/MDM2 protein involved in anthocyanin biosynthesis [272,273]. In comparison to their sensitive counterparts, drought-tolerant wheat and barley types were found to have higher levels of proteins, such as several chaperons (HSP70, HSP90, and cyclophilin A), glutathione-S-transferase (GST), and lipoxygenase (LOX) [274]. Another study found that drought [275], low temperature, and salt [276] caused reductions in RubisCo, enzymes like phosphoglycerokinase (PGK), and transketolase phosphoribulokinase (PRK) in wheat [277]. An increase in numerous minor heat shock proteins, as well as HSP82 from the HSP90 family, was found inside the endosperm of growing wheat seeds exposed to a heat phase [278].
Proteomic investigation and physiological responses in two drought-resistant maize cultivars demonstrated that HSP plays a substantial role in protecting plants from drought stress in a manner [279]. Recently, Halder et al. [280] summarised the proteomic research on salinity, drought stress tolerance, and root system architecture carried out in the previous ten years. They also examined the function of proteomics for abiotic stress tolerance in wheat. It is useful to characterise the proteome of tissues with pathogen infections using proteomic studies of biotic stress. Numerous potent metabolites that are responsible for resilience have been identified in a thorough analysis of the worldwide proteomics studies looking into the reactions of rice to biotic stress [265]. In flax, this area has recently begun to be studied [269]. Some of the examples are given in the Table 7. Table 7. Important genes in flax that are activated and deactivated in response to biotic and abiotic stressors.

Ionomics
Ionomics is the study of the elemental composition of non-metals, metals, and metalloids in various plant species, with a focus on high-throughput detection and quantification [196]. Different plant varieties' ionomic compositions may be determined using high-throughput methods like mass spectrometry with inductive and inductively coupled plasma-atomic emission spectroscopy. It plays a crucial part in helping us comprehend the various elements, their chemical and physical makeup, and how they affect plant metabolism, nourishment, and biochemistry. Due to the diversity of soil types and various edaphic variables affecting growth, plants have developed a range of element uptake skills [286]. Additionally, transit, environmental stress, root uptake capacity, and element abundance all have an impact on the ionomic makeup of a crop.
The quantity of accumulated Na+ and metabolites implicated in glycolysis and the tricarboxylic acid (TCA) cycle were significantly negatively correlated, according to the ionome of wild and cultivated barley that was exposed to various degrees of salt tolerance [287]. Studies on wheat [288], as well as other grasses like Aneurolepidium chinense [289], Setaria viridis [290], and flax, revealed that the concentration of Na + rises with increasing alkalinity stress because plants store significant amounts of Na + in their vacuoles to lower cell water potential. K + , Na + , Ca 2+ , and Cl were the major inorganic ions engaged in osmotic adjustment under PEG-induced water stress in flax, improving drought resilience [291]. In the caseof flax, there were no appreciable variations between the impacts of salt and alkalinity stress on the concentrations of Na + and K + in shoots [292]. This indicates that, unlike other plants like wheat and Chloris virgata, where the K + concentration of shoots was found to be reduced under alkaline stress, flax shoots may have a different adaptive mechanism to alkaline stress. Another study on flax revealed that under salt stress, the amounts of NO 3 in flax shoots dropped, while the uptake of Cl and H 2 PO 4 increased significantly. Therefore, knowing the elemental makeup can help with understanding the process of stress endurance. Ionomics research in flax has not received much notice [196].

Phenomics
High-throughput analysis is used to research phenomics, which is the group of phenotypes combining GxPxE relationships under environmental circumstances [190]. As a result, the phenotype offers the strongest link between surroundings and plant genetics. While phenotypic characterisation has advanced more slowly over the past ten years due to improvements in sequencing technologies, this has limited the discovery of quantitative characteristics, especially those linked to stress resistance [293].
Phenotyping in reaction to abiotic stress remains a significant task because of the complex biosynthetic processes that handle the response of plants to external stressors [294]. Because genomic approaches like GWAS, GS, and QTL rely on high-throughput phenotyping for the improvement of specific traits, the significance of accurate phenotyping has increased in the postgenomic age [295]. The most promise exists for plant breeding when phenomics is used in conjunction with other omic methods. As a result, non-invasive technologies such as magnetic resonance imaging (MRI), lidar (which includes RBG digital imaging) to evaluate growth parameters, and colour imaging of biomass have all been researched to determine the elevated canopy and concealed half (root system) of plants [296,297].
Examples include employing ground-filled rhizoboxes to analyse the architecture of wheat roots using RGB imaging [298] and using RGB digital imaging to phenotype plant shoots [299]. Infrared thermography to confirm the function of stomatal electrical conductance in barley and wheat saplings under salt stress is another illustration [299]. Nuclear magnetic resonance (NMR), computed tomography (CT), and X-rays have all been employed forin situ 3D imaging of root anatomy. There are not many web databases that can help people with image processing, like http://www.plant-image-analysis.org, accessed on 5 May 2023. In order to obtain real-time phenome responses to diseases, the exterior environment, and nutrients, contemporary instruments include hyperspectral imaging systems, fluorescence imaging systems, high-throughput advanced plant phenotyping platforms, laser scanners, and high-resolution IR/NIR cameras. However, to extract phenome data from the enormous databases produced by phenotyping, deep learning techniques are required. Additionally, the thorough administration of platforms and software presents significant difficulties, restricting this application to a few key commodities, like rice, maize, and wheat [196].

Transgenic Flax/Linseed
The initial genetic modification made to flax washerbicide tolerance. The herbicide glyphosate (Roundup) blocks the enzyme 5-enolpyruvylshikimate-3-phosphate (EPSP) polymerase in higher plants. To create transgenic flax lines resistant to 5 mM glyphosate, the glyphosate-insensitive EPSP enzyme from Petunia hybrida was transformed into flax by an Agrobacterium-mediated method of transformation [300]. Later, flax was given additional herbicide-resistant genes, including those for glufosinate ammonium [301] and chlorsulfuron [302]. Under the direction of the CaMV 35S promoter, the phosphinothricin acetyltransferase (PAT) gene from Streptomyces viridochromo was overexpressed. The PAT enzyme was introduced into flax plants to provide them with resistance to the herbicide glufosinate ammonium [301].
To tolerate abiotic stress conditions, several crop plants have undergone genetic modification. Numerous candidate genes, including functional and regulatory genes, have been modified [302]. Functional genes, such as those that encode water channel proteins, ion transporters, late embryogenesis abundant (LEA) protein, and heat shock protein (HSP), are directly engaged in the defence of cells against stressors. The regulatory genes control signal transmission and gene expression during stress reactions, and they comprise transcription factors, protein phosphatases, and protein kinases [303,304]. These studies aiming to genetically modify flax for resistance/tolerance to abiotic stress have not yet been published. When plants are subjected to abiotic stress, the activity of the transporters is first impacted, which hinders the plants' physiological function. The transport of water and other minor solutes, including urea, ammonia, glycerol, silicic acid, boric acid, CO 2 , and H 2 O 2 , is significantly aided by aquaporins in plants [305]. Abiotic stress resistance in various plant species has recently been effectively demonstrated by the application of genes expressing aquaporins. Aquaporins have been discovered in a variety of agricultural plants, including flax, due to their significance [306,307]. A total of 51 AQPs from various subfamilies were found in the flax genome [210].

Conclusions
The production of oilseed crops is substantially affected byan array of abiotic and biotic stressors. To better understand the chemical complexes and biochemical mechanisms involved in plants' ability to tolerate stress, large amounts of data are quickly being created and annotated on a global scale. The availability of various genomic resources in flax has dramatically grown over the past ten years. These resources include molecular markers, linkage maps, transcriptomes, and whole-genome sequencing. Through a varietal development programme, these resources may be effectively used to increase the capacity of flax for adaptation to the environment and resistance to biotic stress. Given its great economic worth, flax is used in a variety of ways in the food, bioenergy, nutrition, and nutraceutical industries. This review has covered many aspects of tolerance that are now being utilised to overcome these barriers. These strategies really work to increase a plant's natural ability to withstand diverse biotic and abiotic stresses, possibly having long-lasting impacts and, in theory, not negatively affecting other intriguing agricultural attributes like yield and the quality of edible parts. Additionally, technological advancements like the application of high-throughput platforms to measure the tolerance traits of crop genotypes upon treatments give researchers the chance to characterise the plant responses of a subset of varieties under stringent growing circumstances or in the field, where real-world agricultural conditions occur.

Conflicts of Interest:
The authors declare no conflict of interest.