Characterization for Drought Tolerance and Physiological Efficiency in Novel Cytoplasmic Male Sterile Sources of Sunflower (Helianthus annuus L.)

Abstract: Sunflower is sensitive to drought and its hybrids have a limited cytoplasmic diversity. The wild 10 cytoplasmic sources of sunflower are not well exploited to their potential for drought tolerance and hybrid 11 development. In this respect, we carried out a Line × Tester based genetic study using 19 sunflower 12 genotypes representing, 13 cytoplasmic male sterile (CMS) lines from wild and conventional sources, 2 13 maintainer lines, and 4 restorer lines. The CMS and maintainer lines were crossed with restorer lines to 14 develop sixty one-way F1 hybrids. The parents and their hybrids were evaluated under two water 15 regimes viz., normal irrigated and water stress. A total of twelve important plant descriptors were studied 16 over a period of two years. The significant differences were observed between parents and hybrids in both 17 water regimes. Hybrids were higher in average values for all the descriptors than parents. The role of 18 female parent was more prominent in the expression of traits in hybrids as compared to male parents. The 19 CMS sources varied significantly regarding seed yield per plant and other physiological traits. Proline 20 content was three times higher in parents and their hybrids under water stress, and it was not correlated 21 with any other descriptor. Accession CMS-PKU-2A was identified as the best general combiner for leaf 22 area and specific leaf weight. Whereas, CMS-234A was the best general combiner for biological yield and 23 photosynthetic efficiency under both the conditions. The cross combinations CMS-ARG-2A × RCR-8297, 24 CMS-234A × P124R, and CMS-38A × P124R were found significant for biological yield, seed yield and oil 25 content under both environments. Overall, this study provides useful information about the cytoplasmic 26 effects on important sunflower traits and drought stress tolerance when used in the different 27 combinations. 28


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their hybrid combinations performed better under normal conditions than in the stress environment (Table   83 1). Likewise, hybrids were with more leaves, higher harvest index, seed yield, and oil content. Whereas, 84 under water stress, parents and hybrids overlapped for harvest index and proline content (Table 1). The 85 proline content was almost three times more in water-limited filed. Whereas, the photosynthetic efficiency 86 remained to be unaltered under normal and drought condition (Table 1)

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The cophenetic correlation coefficient of clustering is 0.8.

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. CC-BY 4.0 International license available under a not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made The copyright holder for this preprint (which was this version posted August 26, 2018. ; https://doi.org/10.1101/400531 doi: bioRxiv preprint . CC-BY 4.0 International license available under a not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made The analysis of variance for combining abilities of the twelve descriptors studied in a line x tester

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(15 x 4) design is presented in Table 2. The mean squares due to treatments were highly significant for all 96 the traits except for the proline content (Table 2). Likewise, the mean squares due to lines (female), testers 97 (males), and female x male interactions were recorded highly significant for all the traits under both the 98 environments and also, over the years (Table 2). Parentals and their hybrids had a highly significant (P ≤ 99 0.01) general combining ability (GCA) and specific combining ability (SCA) effects, for all the traits and under 100 both the environments ( Table 2). The ratios of GCA/SCA effect were >0.5 for biological yield and oil content 101 under normal water environment, while under water stress environment leaf water potential, 102 photosynthetic efficiency and harvest index, suggesting the predominance of additive over non-additive 103 genetic effects. This ratio was <0.5 for most of the traits studied in both environments, implying a significant 104 role of non-additive genetic effect on these traits (Table 2).

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The proportional contribution of parent and their interactions, the contribution of female parents 106 were observed to be higher compared to testers (males) irrespective of the environments (Table 2).

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However, their overall interaction component (lines x testers) had a higher proportional contribution for 108 traits viz., leaf area, specific leaf weight, leaf area index, leaf water potential, relative leaf water content, 109 proline content, seed yield, harvest index and oil content (Table 2).

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In case of specific leaf weight, CMS-PKU-2A recorded highly significant GCA under normal as well as in 24 water stress environment ( Table 3). The male parent P100R was observed as a good combiner for this trait 25 under both the environments. A total of eighteen hybrids recorded highly significant positive SCA effects 26 under both the environments (Table S1). Whereas, the top three common combinations for SCA were 27 CMS-ARG-2A × RCR-8297, CMS-XA × P69R, and CMS-234A × P100R (Table 4).

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Among male parents, P100R was observed with a significant GCA effect (0.19 and 0.21) under both 32 environments (

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For leaf water potential under the normal environment, CMS-ARG-3A was observed as significant 37 combiner (Table 3)

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CMS-38A and NC-41B were observed as better general combiners for leaf water potential (Table 3). A total 39 of twenty-one hybrids were reported to have significant positive SCA effects under normal environment 40 whereas, twenty-seven hybrids had the good combining ability under water stress environment (Table S1).

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The best three cross combinations identified with high SCA effects for leaf water potential under both the 42 . CC-BY 4.0 International license available under a not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made

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The six CMS analogues out of nine and both the maintainers were recorded as significant 46 general combiners for relative leaf water content were identified as good combiners under the normal 47 environment (Table 3). In water stress environment, female lines CMA-XA, CMS-PKU-2A, CMS-ARG-2A and 48 CMS-PHIR-27A from wild sources and CMS-NC-41B from the PET-1 source were observed as significant 49 combiners (Table 3). A total of twenty-eight hybrids recorded significant positive SCA effects in the normal 50 environment. Twenty-nine hybrids recorded significant positive SCA effects under stress environment (Table   51   S1). The three cross combinations viz. CMS-234A × RCR-8297, CMS-E002-91A × P69R, and CMS-40A × P69R 52 were identified with highly significant SCA effects for relative leaf water content under both the 53 environments (Table 4).

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In the normal environment and water stress environment, the female parents CMS-234A was observed 56 with the highest GCA effect (5.56 and 4.68 1) ( Table 3). While the male parent RCR-8297 and P69R were 57 recorded as a significant combiners (0.93 and 1.34; 1.67 and 0.75) under both the environments for 58 photosynthetic efficiency (Table 3). Twenty-one cross combinations recorded significant positive SCA 59 effects in a normal environment. In the water stress condition, seventeen cross combinations were 60 identified with high SCA values (Table S1)

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(3.39) (from PET-1) were recorded as significant for the GCA effects (Table 3) (Table S1).  (Table   87   . CC-BY 4.0 International license available under a not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made

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All CMS analogues from wild sources were recorded as very good combiner except CMS-ARG-6A 92 whereas, from conventional PET-1 source, none of the female lines had significant GCA for harvest index 93 under water stress environment ( Table 3). The tester RCR-8297 appeared to be a good general combiner for 94 harvest index with significant and positive GCA effects (2.54 and 2.88) under both the environments (Table   95 3). Twenty-four hybrids under normal environment and twenty hybrids under water stress environment 96 recorded significant positive SCA effects (Table S1). Best three hybrids were CMS-PHIR-27A × RCR-8297, 42B 97 × P100R and CMS-ARG-3A × P124R for significant SCA effects for harvest index common to both 98 environments (Table 4).  (Table 3). Whereas, in 103 water stress environment, all CMS analogues except CMS-ARG-6A and CMS-PHIR-27A were observed 104 significant combiner for oil content ( Table 3). The male parent RCR-8297 and P69R were recorded as highly 105 significant general combiner under stress environment, while P100R was recorded as a significant general 106 combiner under the normal environment (Table 3). Twenty-two hybrids under normal environment and 107 twenty-eight hybrids under stress environment recorded significant positive SCA effects (Table S1).

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In case of normal environment, 16 correlation coefficients were found to be significant (P < 0.05) (Figure 2).

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Among them four were negative correlations and three were absolute correlations. Harvest index was found 116 negatively correlated with biological yield, specific leaf weight, leaf area leaf area index (Figure 2). While 117 absolute correlations exist between leaf area and leaf area index. Also, specific leaf weight with leaf area 118 and leaf area index (0.92) (Figure 2). Whereas, under water stress environment, a total of 14 correlation 119 coefficients were found to be significant (P < 0.05) (Figure 4). Thirteen were positive correlations and one 120 was a negative correlation i.e. between harvest index and biological yield (-0.59) (Figure 3). Also, the 121 specific leaf weight was absolutely correlated with leaf area (0.90) and leaf area index (0.92) (Figure 3). The            (Table 5). Both, A lines (13) and 205 B lines (2) were crossed with the R lines (4) to produce sixty one-way F1 hybrids (Table 5).
The hybrids 206 were produced as the nine alloplasmic lines and euplasmic CMS lines from petiolaris source were crossed 207 with restorers to synthesize a set of 52 A × R crosses (TABLE 5)  Week 1 Week 2 Week 3 Week 4 Week 5 Week 6 Week 7 Week 8 Week 9 Week 10 Week 11 Week 12 Week 13 Week 14 Week 15 Week 16 Week 17 Week 18 Week 19 Week 20 Temperature ( Physical properties of soil were also analyzed to determine the native fertility and soil texture of 235 the experimental fields (

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Whereas, the specific leaf weight (g) was estimated as total leaf weight divided by the number of leaves. The

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. CC-BY 4.0 International license available under a not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made The copyright holder for this preprint (which was this version posted August 26, 2018. ; https://doi.org/10.1101/400531 doi: bioRxiv preprint