Identification and Characterization of Salt-Responsive MicroRNAs in Taxodium hybrid ‘Zhongshanshan 405’ by High-Throughput Sequencing
Abstract
:1. Introduction
2. Materials and Methods
2.1. Plant Materials
2.2. Small RNA Library Construction and High-Throughput Sequencing
2.3. Data Filtering and Mapping Reads
2.4. Identification of Known MicroRNAs and Novel MicroRNAs
2.5. Analyzing sRNA Expression
2.6. Target Prediction
2.7. Validation of miRNAs’ Expression by Real-Time Quantitative PCR (QRT-PCR)
3. Results
3.1. Deep-Sequencing of sRNAs
3.2. Identification of Known miRNAs and Novel miRNAs
3.3. Target Prediction for Known and Novel miRNAs
3.4. QRT-PCR Validation
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Data Availability Statement
Conflicts of Interest
References
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Sample | Total Reads | N% > 10% | Low Quality | 5 Adapter Contamine | 3 Adapter Null or Insert Null | with ployA/T/G/C | Clean Reads |
---|---|---|---|---|---|---|---|
T1-1 | 13,212,250 (100.00%) | 34 (0.00%) | 10,535 (0.08%) | 10,746 (0.08%) | 419,301 (3.17%) | 11,549 (0.09%) | 12,760,085 (96.58%) |
T1-2 | 14,554,682 (100.00%) | 34 (0.00%) | 10,988 (0.08%) | 15,718 (0.11%) | 422,272 (2.90%) | 16,198 (0.11%) | 14,089,472 (96.80%) |
T1-3 | 11,047,511 (100.00%) | 33 (0.00%) | 6373 (0.06%) | 13,511 (0.12%) | 371,044 (3.36%) | 11,949 (0.11%) | 10,644,601 (96.35%) |
T2-1 | 10,616,411 (100.00%) | 21 (0.00%) | 4386 (0.04%) | 17,123 (0.16%) | 269,497 (2.54%) | 19,629 (0.18%) | 10,305,755 (97.07%) |
T2-2 | 14,511,597 (100.00%) | 33 (0.00%) | 7570 (0.05%) | 20,556 (0.14%) | 450,817 (3.11%) | 20,197 (0.14%) | 14,012,424 (96.56%) |
T2-3 | 13,778,931 (100.00%) | 32 (0.00%) | 7794 (0.06%) | 17,722 (0.13%) | 784,585 (5.69%) | 30,837 (0.22%) | 12,937,961 (93.90%) |
T3-1 | 12,036,280 (100.00%) | 28 (0.00%) | 7326 (0.06%) | 8488 (0.07%) | 369,020 (3.07%) | 18,987 (0.16%) | 11,632,431 (96.64%) |
T3-2 | 10,291,894 (100.00%) | 10 (0.00%) | 4139 (0.04%) | 11,050 (0.11%) | 455,849 (4.43%) | 12,135 (0.12%) | 9,808,711 (95.31%) |
T3-3 | 11,345,585 (100.00%) | 26 (0.00%) | 4628 (0.04%) | 6330 (0.06%) | 348,991 (3.08%) | 7011 (0.06%) | 10,978,599 (96.77%) |
T4-1 | 11,909,066 (100.00%) | 36 (0.00%) | 5711 (0.05%) | 8714 (0.07%) | 407,633 (3.42%) | 10,874 (0.09%) | 11,476,098 (96.36%) |
T4-2 | 12,833,399 (100.00%) | 32 (0.00%) | 6067 (0.05%) | 15,353 (0.12%) | 519,695 (4.05%) | 8185 (0.06%) | 12,284,067 (95.72%) |
T4-3 | 12,115,728 (100.00%) | 35 (0.00%) | 8663 (0.07%) | 9867 (0.08%) | 474,255 (3.91%) | 14,847 (0.12%) | 11,608,061 (95.81%) |
Types | Total | T1-1 | T1-2 | T1-3 | T2-1 | T2-2 | T2-3 | T3-1 | T3-2 | T3-3 | T4-1 | T4-2 | T4-3 | |
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Mapped known miRNAs | Mature | 49 | 36 | 43 | 39 | 31 | 30 | 29 | 40 | 26 | 29 | 19 | 31 | 26 |
Hairpin | 55 | 38 | 48 | 44 | 34 | 34 | 32 | 43 | 31 | 33 | 22 | 36 | 28 | |
Mapped novel miRNAs | Mature | 98 | 88 | 88 | 83 | 77 | 74 | 72 | 80 | 62 | 77 | 57 | 73 | 70 |
Hairpin | 100 | 92 | 93 | 88 | 84 | 80 | 79 | 85 | 67 | 83 | 66 | 81 | 76 | |
Star | 68 | 44 | 42 | 40 | 37 | 25 | 30 | 39 | 17 | 40 | 16 | 32 | 18 |
miRNA | Target | Expectation | Target Accessibility | Target Description | Inhibition | Multiplicity | |
---|---|---|---|---|---|---|---|
T1 vs. T2 | novel_118 | CL16860Contig1 | 4.5 | 18.248 | Putative polyprotein | Cleavage | 1 |
novel_13 | CL15Contig4 | 4.5 | 20.381 | Probable disease resistance protein | Translation | 1 | |
novel_16 | T3_Unigene_BMK.33047 | 4 | 18.583 | Putative truncated TIR-NBS-LRR protein | Translation | 1 | |
novel_52 | CL10744Contig1 | 4.5 | 10.073 | Zinc finger CCCH domain-containing protein 35 | Translation | 1 | |
novel_78 | CL25843Contig1 | 4.5 | 12.165 | Probable nucleoredoxin 1 | Translation | 1 | |
miR160a | CL8543Contig1 | 4 | 5.964 | Chaperone protein dnaJ 11 | Translation | 1 | |
miR396b | CL5428Contig1 | 3.5 | 22.527 | Glycosyltransferase family protein 2 | Cleavage | 1 | |
T1 vs. T3 | novel_123 | CL27539Contig1 | 4.5 | 13.132 | RNA-binding protein 25 | Cleavage | 1 |
novel_21 | CL18504Contig1 | 4 | 24.442 | TMV resistance protein N | Translation | 1 | |
novel_41 | CL2111Contig1 | 5 | 12.456 | Mitogen-activated protein kinase kinase kinase | Cleavage | 1 | |
novel_4 | CL1685Contig1 | 4.5 | 12.818 | Ethylene-responsive transcription factor RAP2-13 | Cleavage | 1 | |
novel_77 | T2_Unigene_BMK.14386 | 4.5 | 16.719 | G-type lectin S-receptor-like serine/threonine-protein kinase | Cleavage | 1 | |
miR156a | CL14355Contig1 | 4 | 18.821 | RNA and export factor-binding protein 2 | Cleavage | 1 | |
miR319a | CL11314Contig1 | 4 | 17.74 | Beta-amylase 1 isoform 1 | Cleavage | 1 | |
T1 vs. T4 | novel_100 | CL24684Contig1 | 4.5 | 17.879 | ATP synthase subunit | Cleavage | 1 |
novel_13 | CL4989Contig1 | 4 | 20.269 | Salicylate O-methyltransferase | Cleavage | 1 | |
novel_14 | CL1013Contig1 | 3.5 | 18.997 | Probable LRR receptor-like serine/threonine-protein kinase | Cleavage | 1 | |
novel_29 | CL11748Contig1 | 2 | 18.906 | Glycerol-3-phosphate 2-O-acyltransferase 6 | Translation | 1 | |
novel_2 | CL14285Contig1 | 4 | 16.055 | G-type lectin S-receptor-like serine/threonine-protein kinase | Translation | 1 | |
novel_40 | CL1182Contig1 | 5 | 18.447 | Disease resistance RPP13-like protein 4 | Translation | 1 | |
novel_41 | CL2111Contig1 | 5 | 12.456 | Mitogen-activated protein kinase kinase kinase | Cleavage | 1 | |
novel_42 | CL1146Contig1 | 2.5 | 13.999 | F-box/LRR-repeat protein 17 | Cleavage | 1 | |
novel_77 | CL1110Contig2 | 4 | 19.906 | Cysteine-rich receptor-like protein kinase | Cleavage | 1 | |
novel_98 | CL13461Contig1 | 4 | 14.38 | Homeobox-leucine zipper protein ATHB-13 | Cleavage | 1 | |
miR159a | CL12428Contig1 | 3 | 16.021 | Chlorophyll a-b binding protein 7 | Translation | 1 | |
miR396a-5p | CL10009Contig1 | 5 | 20.004 | DNA replication licensing factor mcm5 | Cleavage | 1 | |
miR396f | CL2465Contig1 | 4 | 18.647 | U-box domain-containing protein 12 | Cleavage | 1 | |
miR399d | CL1025Contig1 | 5 | 20.819 | Tonoplast dicarboxylate transporter | Cleavage | 1 | |
T2 vs. T3 | novel_100 | CL772Contig3 | 4.5 | 13.205 | Transcription factor MYB59 | Cleavage | 1 |
novel_108 | CL805Contig1 | 4 | 14.766 | LRR receptor-like serine/threonine-protein kinase | Translation | 1 | |
novel_111 | CL1347Contig2 | 3.5 | 14.988 | Subtilisin-like protease | Cleavage | 1 | |
novel_123 | CL14243Contig1 | 4.5 | 12.566 | Trehalose-phosphatase | Translation | 1 | |
novel_16 | CL22Contig4 | 5 | 21.524 | TMV resistance protein | Translation | 1 | |
novel_24 | CL228Contig1 | 2.5 | 20.215 | TMV resistance protein | Cleavage | 1 | |
novel_30 | CL14581Contig1 | 4.5 | 19.087 | Xyloglucan endotransglucosylase/hydrolase | Translation | 1 | |
novel_41 | CL23589Contig1 | 4.5 | 13.732 | Chaperone protein dnaJ | Cleavage | 1 | |
novel_52 | T3_Unigene_BMK.32994 | 4.5 | 23.774 | Protein LURP-one-related | Cleavage | 1 | |
novel_77 | T2_Unigene_BMK.14386 | 4.5 | 16.719 | G-type lectin S-receptor-like serine/threonine-protein kinase | Cleavage | 1 | |
novel_78 | CL10767Contig1 | 5 | 17.257 | BON1-associated protein | Translation | 1 | |
novel_88 | CL15264Contig1 | 4 | 14.363 | Cysteine-rich receptor-like protein kinase | Translation | 1 | |
novel_89 | T3_Unigene_BMK.16696 | 3 | 17.651 | Squamosa promoter-binding-like protein | Cleavage | 1 | |
pab-miR159a | CL2378Contig1 | 3 | 16.57 | Cinnamoyl CoA reductase | Translation | 1 | |
pab-miR319a | CL26045Contig1 | 4 | 17.17 | Disease resistance protein RPS2 | Cleavage | 1 | |
T3 vs. T4 | novel_108 | CL1146Contig1 | 4 | 13.459 | F-box/LRR-repeat protein 17 | Translation | 1 |
novel_13 | CL4989Contig1 | 4 | 20.269 | Salicylate O-methyltransferase | Cleavage | 1 | |
novel_14 | CL1013Contig1 | 3.5 | 18.997 | Probable LRR receptor-like serine/threonine-protein kinase | Cleavage | 1 | |
miR156a | CL24146Contig1 | 3 | 15.592 | Probable LRR receptor-like serine/threonine-protein kinase | Cleavage | 1 | |
miR159a | CL12428Contig1 | 3 | 16.021 | Chlorophyll a-b binding protein 7 | Translation | 1 | |
miR396a-5p | CL10009Contig1 | 5 | 20.004 | DNA replication licensing factor mcm5 | Cleavage | 1 | |
miR396b | CL2465Contig1 | 4.5 | 18.647 | U-box domain-containing protein 12 | Cleavage | 1 | |
miR396f | CL13812Contig1 | 4.5 | 16.915 | Potassium transporter 1 | Translation | 1 |
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Wang, Z.; Zhang, F.; Shi, Q.; Zhang, R.; Yin, Y.; Yu, C. Identification and Characterization of Salt-Responsive MicroRNAs in Taxodium hybrid ‘Zhongshanshan 405’ by High-Throughput Sequencing. Forests 2022, 13, 1685. https://doi.org/10.3390/f13101685
Wang Z, Zhang F, Shi Q, Zhang R, Yin Y, Yu C. Identification and Characterization of Salt-Responsive MicroRNAs in Taxodium hybrid ‘Zhongshanshan 405’ by High-Throughput Sequencing. Forests. 2022; 13(10):1685. https://doi.org/10.3390/f13101685
Chicago/Turabian StyleWang, Zhiquan, Fengjiao Zhang, Qin Shi, Rui Zhang, Yunlong Yin, and Chaoguang Yu. 2022. "Identification and Characterization of Salt-Responsive MicroRNAs in Taxodium hybrid ‘Zhongshanshan 405’ by High-Throughput Sequencing" Forests 13, no. 10: 1685. https://doi.org/10.3390/f13101685
APA StyleWang, Z., Zhang, F., Shi, Q., Zhang, R., Yin, Y., & Yu, C. (2022). Identification and Characterization of Salt-Responsive MicroRNAs in Taxodium hybrid ‘Zhongshanshan 405’ by High-Throughput Sequencing. Forests, 13(10), 1685. https://doi.org/10.3390/f13101685