Xinjiang Agricultural Sciences ›› 2021, Vol. 58 ›› Issue (12): 2265-2273.DOI: 10.6048/j.issn.1001-4330.2021.12.014
• Plant Protection· Horticultural Special Local Products· Soil Fertilizer· Water Saving Irrigation Agroecological Environment· Agricultural Equipment Engineering and Mechanization • Previous Articles Next Articles
LING Yueming, LI Meihua, YANG Yong, YANG Wenli, FAN Rong, YI Hongping, ZHANG Hong, ZHANG Xuejun
Received:
2021-01-30
Online:
2021-12-20
Published:
2021-12-31
Supported by:
凌悦铭, 李寐华, 杨永, 杨文莉, 范蓉, 伊鸿平, 张红, 张学军
基金资助:
CLC Number:
LING Yueming, LI Meihua, YANG Yong, YANG Wenli, FAN Rong, YI Hongping, ZHANG Hong, ZHANG Xuejun. Development of InDel Marker for Melon Resistance to Downy Mildew Based on BSA Resequencing[J]. Xinjiang Agricultural Sciences, 2021, 58(12): 2265-2273.
凌悦铭, 李寐华, 杨永, 杨文莉, 范蓉, 伊鸿平, 张红, 张学军. 基于BSA-Seq技术的甜瓜抗霜霉病InDel标记开发[J]. 新疆农业科学, 2021, 58(12): 2265-2273.
样本名 Sample | 原始数据量 Raw Base (bp) | 有效数据量 Clean Base (bp) | 碱基错误率 Error Rate(%) | Q20(%) | Q30(%) | GC含量 GC Content(%) |
---|---|---|---|---|---|---|
DM-4 | 5 230 484 100 | 5 205 287 100 | 0.01 | 97.74 | 94.94 | 37.39 |
DM-2 | 5 954 570 100 | 5 926 054 500 | 0.01 | 97.83 | 95.10 | 37.23 |
DM-R | 9 302 519 100 | 9 266 289 000 | 0.01 | 97.93 | 95.27 | 37.17 |
DM-S | 9 931 454 700 | 9 878 084 400 | 0.01 | 97.81 | 95.08 | 37.12 |
Table 1 The quality of sequencing data
样本名 Sample | 原始数据量 Raw Base (bp) | 有效数据量 Clean Base (bp) | 碱基错误率 Error Rate(%) | Q20(%) | Q30(%) | GC含量 GC Content(%) |
---|---|---|---|---|---|---|
DM-4 | 5 230 484 100 | 5 205 287 100 | 0.01 | 97.74 | 94.94 | 37.39 |
DM-2 | 5 954 570 100 | 5 926 054 500 | 0.01 | 97.83 | 95.10 | 37.23 |
DM-R | 9 302 519 100 | 9 266 289 000 | 0.01 | 97.93 | 95.27 | 37.17 |
DM-S | 9 931 454 700 | 9 878 084 400 | 0.01 | 97.81 | 95.08 | 37.12 |
样本名 Sample | 比对序列数 Mapped reads | 有效序列数 Total reads | 比对率 Mapping rate(%) | 平均测序深度 Average depth(X) | 1X覆盖度 Coverage at least 1X(%) |
---|---|---|---|---|---|
DM-2 | 36 714 906 | 39 507 030 | 92.93 | 13.11 | 96.38 |
DM-4 | 32 024 597 | 34 701 914 | 92.28 | 11.57 | 96.37 |
DM-S | 61 510 228 | 65 853 896 | 93.40 | 21.29 | 97.75 |
DM-R | 57 254 273 | 61 775 260 | 92.68 | 19.85 | 97.70 |
Table 2 Sequencing depth and coverage
样本名 Sample | 比对序列数 Mapped reads | 有效序列数 Total reads | 比对率 Mapping rate(%) | 平均测序深度 Average depth(X) | 1X覆盖度 Coverage at least 1X(%) |
---|---|---|---|---|---|
DM-2 | 36 714 906 | 39 507 030 | 92.93 | 13.11 | 96.38 |
DM-4 | 32 024 597 | 34 701 914 | 92.28 | 11.57 | 96.37 |
DM-S | 61 510 228 | 65 853 896 | 93.40 | 21.29 | 97.75 |
DM-R | 57 254 273 | 61 775 260 | 92.68 | 19.85 | 97.70 |
Fig. 2 Distribution curve of Δ( SNP-Index ) Note: Orange curve represents the confidence threshold line of 0.95, and green curve represents the confidence threshold line of 0.99
Fig. 3 Polymorphism detected by InDel primers among F2 individual plants Note: M:Marker I; P1: PCR amplification in DM-4; P2: PCR amplification in DM-2
名称 Name | 正向引物序列5’- 3’ Forward primers sequence 5’ - 3’ | 反向引物5’- 3’ Reverse primers sequence 5’ - 3’ | 是否具有多态性 Polymorphism or not |
---|---|---|---|
InDel 1 | AGGTAAAGCCCCAGAGAGGA | CCTCCCGTGAAGTTTGGAGT | 否 |
InDel 2 | AGTGAATGTGGTCATTGTGCT | CCTCCCGTGAAGTTTGGAGT | 是 |
InDel 3 | ACACAGCACAACCCAGACAA | CTACGACCAACCCCCACATG | 是 |
InDel 4 | GCACTCTCCAGCCTTGAACT | GAGCAAAGGTACCGCTGTCT | 是 |
InDel 5 | TTTTCATTCGGATTCCTTGTTGT | AAGCTATGAGACCTACGCGC | 否 |
InDel 6 | TCAAAGTGACCCCATCCAGTG | TGTTTTGAGTTCAACGGGGGA | 否 |
InDel 7 | GTGACCCACGATAGGTACACC | TCAAGTTTGGGAGATTGGATCA | 否 |
InDel 8 | AGCATTCTCCCAGTTTCCCC | TGGGAACAAGAAAGTCAGGCA | 否 |
InDel 9 | ACACAATGAACCAAGATATTTGCCA | TGTTTGTAAGCCACTCGACT | 是 |
InDel 10 | ACGACACTAAATGACAGTCCGT | CCAAAATGGAAACCCATGCCC | 否 |
InDel 11 | GCAAAGTTGACCCTCTGTTCG | CAGCGTTCTAGGGGTTGGTT | 否 |
InDel 12 | TGTGAGAATGACAGCCCACA | CACATCTCTCAAGAGGCGGA | 否 |
InDel 13 | CCTCGCTCTATTTGCATTCACA | TTTATACGCGTACTAGTTCTCTCA | 否 |
InDel 14 | AAAGTCAGGATGGCCGAGTG | TCTCTTTCATCAGCGGCGTT | 是 |
InDel 15 | TCACAGCTCACAATCACAGGT | AGGGGAAGCTGGAGAAGACA | 是 |
InDel 16 | ACCTGCCGTCTATCGTGTTT | TGAGGTCGGTCCAAACAAGT | 否 |
InDel 17 | GCTGCCGTTTCGTTTTCAGT | ACGACCAAGAAACCCAGGAA | 否 |
InDel 18 | CCGTTGATTGCGCTAACACA | TGCCGCACGAGTATTCACTT | 是 |
InDel 19 | AGGAGTTTGGGTATGACACGAC | TTCTCCCTTTACCCGTTGCC | 是 |
InDel 20 | TGGGGTTGCTTGTGGATAGT | AGGCGAACACCTTTTGATTCA | 是 |
InDel 21 | GGAAGAGGAAGAGGCAGAGG | ACGAATCATTTCCTCCTCCGA | 否 |
InDel 22 | TGCTCTTGAGGCTAGGGCTA | TAGAGAGGTCCGAGCCCATC | 否 |
InDel 23 | TCATTGGCTAGAACTTCGACCA | CGAAAGTGATGGCCATGGGA | 否 |
InDel 24 | AGTGGTTGGTGGTGCATAGG | GCAACACTTTGATCCCCACA | 否 |
InDel 25 | ACCTCCAAGAAATGAATTCAGGA | TGAACGACGACGACCAATCT | 否 |
InDel 26 | CTTGCAAGGGCTAATGGTGC | ACGAATCAAAGGTGTAGCCA | 否 |
InDel 27 | ACAGTTTAGCTTCCATCCCTACA | TCGTTTAGGTCATTGTCCTCCA | 否 |
Table 3 The information of InDel primers
名称 Name | 正向引物序列5’- 3’ Forward primers sequence 5’ - 3’ | 反向引物5’- 3’ Reverse primers sequence 5’ - 3’ | 是否具有多态性 Polymorphism or not |
---|---|---|---|
InDel 1 | AGGTAAAGCCCCAGAGAGGA | CCTCCCGTGAAGTTTGGAGT | 否 |
InDel 2 | AGTGAATGTGGTCATTGTGCT | CCTCCCGTGAAGTTTGGAGT | 是 |
InDel 3 | ACACAGCACAACCCAGACAA | CTACGACCAACCCCCACATG | 是 |
InDel 4 | GCACTCTCCAGCCTTGAACT | GAGCAAAGGTACCGCTGTCT | 是 |
InDel 5 | TTTTCATTCGGATTCCTTGTTGT | AAGCTATGAGACCTACGCGC | 否 |
InDel 6 | TCAAAGTGACCCCATCCAGTG | TGTTTTGAGTTCAACGGGGGA | 否 |
InDel 7 | GTGACCCACGATAGGTACACC | TCAAGTTTGGGAGATTGGATCA | 否 |
InDel 8 | AGCATTCTCCCAGTTTCCCC | TGGGAACAAGAAAGTCAGGCA | 否 |
InDel 9 | ACACAATGAACCAAGATATTTGCCA | TGTTTGTAAGCCACTCGACT | 是 |
InDel 10 | ACGACACTAAATGACAGTCCGT | CCAAAATGGAAACCCATGCCC | 否 |
InDel 11 | GCAAAGTTGACCCTCTGTTCG | CAGCGTTCTAGGGGTTGGTT | 否 |
InDel 12 | TGTGAGAATGACAGCCCACA | CACATCTCTCAAGAGGCGGA | 否 |
InDel 13 | CCTCGCTCTATTTGCATTCACA | TTTATACGCGTACTAGTTCTCTCA | 否 |
InDel 14 | AAAGTCAGGATGGCCGAGTG | TCTCTTTCATCAGCGGCGTT | 是 |
InDel 15 | TCACAGCTCACAATCACAGGT | AGGGGAAGCTGGAGAAGACA | 是 |
InDel 16 | ACCTGCCGTCTATCGTGTTT | TGAGGTCGGTCCAAACAAGT | 否 |
InDel 17 | GCTGCCGTTTCGTTTTCAGT | ACGACCAAGAAACCCAGGAA | 否 |
InDel 18 | CCGTTGATTGCGCTAACACA | TGCCGCACGAGTATTCACTT | 是 |
InDel 19 | AGGAGTTTGGGTATGACACGAC | TTCTCCCTTTACCCGTTGCC | 是 |
InDel 20 | TGGGGTTGCTTGTGGATAGT | AGGCGAACACCTTTTGATTCA | 是 |
InDel 21 | GGAAGAGGAAGAGGCAGAGG | ACGAATCATTTCCTCCTCCGA | 否 |
InDel 22 | TGCTCTTGAGGCTAGGGCTA | TAGAGAGGTCCGAGCCCATC | 否 |
InDel 23 | TCATTGGCTAGAACTTCGACCA | CGAAAGTGATGGCCATGGGA | 否 |
InDel 24 | AGTGGTTGGTGGTGCATAGG | GCAACACTTTGATCCCCACA | 否 |
InDel 25 | ACCTCCAAGAAATGAATTCAGGA | TGAACGACGACGACCAATCT | 否 |
InDel 26 | CTTGCAAGGGCTAATGGTGC | ACGAATCAAAGGTGTAGCCA | 否 |
InDel 27 | ACAGTTTAGCTTCCATCCCTACA | TCGTTTAGGTCATTGTCCTCCA | 否 |
名称 Name | 正向引物序列5’- 3’ Forward primers sequence 5’ - 3’ | 反向引物5’- 3’ Reverse primers sequence 5’ - 3’ | 是否具有多态性 Polymorphism or not |
---|---|---|---|
InDel 28 | TCATTGGCCGACACATGTCA | ACTCCAAAACTCCAAACCCCA | 否 |
InDel 29 | TGGTCCCTAACTAAATTACCCTCA | TCCCATAGCCGCCAACATTT | 否 |
InDel 30 | CTCAACTGGGTGTCGCTTCT | ATCGCAATGCCCCCAGTTAA | 否 |
InDel 31 | AACAGAGCTAACTTTGGCGC | TGTGAGAGTTTGGGCTTCCA | 否 |
InDel 32 | TCGAGCTAAAATGATTGTTGGCA | TCCTTCAAACATAAGGACACTCT | 否 |
InDel 33 | ACGAAACTCAATACTCGAGTGGA | TCACCACCTCCAATTCTTTGT | 否 |
InDel 34 | AAAAGCACCCCTCCCTTAGC | CGTACTCTAGACGTGTGCGG | 否 |
InDel 35 | TCTTTCACCCGTCAACTTAACT | TGTCCCACTTGAGTTGCATT | 否 |
InDel 36 | TGGTTGGGGAGCAAAATACA | AAACGTCCGTCGAAAATGCC | 否 |
InDel 37 | CGGCCGAATGGATGCTTTTT | TGCATTCGTTTTGGTTGGCA | 否 |
Table 3 The information of InDel primers
名称 Name | 正向引物序列5’- 3’ Forward primers sequence 5’ - 3’ | 反向引物5’- 3’ Reverse primers sequence 5’ - 3’ | 是否具有多态性 Polymorphism or not |
---|---|---|---|
InDel 28 | TCATTGGCCGACACATGTCA | ACTCCAAAACTCCAAACCCCA | 否 |
InDel 29 | TGGTCCCTAACTAAATTACCCTCA | TCCCATAGCCGCCAACATTT | 否 |
InDel 30 | CTCAACTGGGTGTCGCTTCT | ATCGCAATGCCCCCAGTTAA | 否 |
InDel 31 | AACAGAGCTAACTTTGGCGC | TGTGAGAGTTTGGGCTTCCA | 否 |
InDel 32 | TCGAGCTAAAATGATTGTTGGCA | TCCTTCAAACATAAGGACACTCT | 否 |
InDel 33 | ACGAAACTCAATACTCGAGTGGA | TCACCACCTCCAATTCTTTGT | 否 |
InDel 34 | AAAAGCACCCCTCCCTTAGC | CGTACTCTAGACGTGTGCGG | 否 |
InDel 35 | TCTTTCACCCGTCAACTTAACT | TGTCCCACTTGAGTTGCATT | 否 |
InDel 36 | TGGTTGGGGAGCAAAATACA | AAACGTCCGTCGAAAATGCC | 否 |
InDel 37 | CGGCCGAATGGATGCTTTTT | TGCATTCGTTTTGGTTGGCA | 否 |
Fig.4 Result of amplification by primers InDel 15, InDel 20 among parents and F2 individual plants Note: Fig.2A: Result of amplification by primers InDel 15. 11: DM-4, 12: DM-2, 1~5: resistant individual, 6~10: susceptible individual. Fig.2B: Result of amplification by primers InDel 20. 2: DM-4, 1: DM-2, 3~7: resistant individual, 8~12: susceptible individual. M: DL2000 DNA Marker
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