基于SLAF-seq技术的甘薯SNP位点开发

doi:10.3864/j.issn.0578-1752.2016.01.003

Abstract

Abstract: 【Objective】 Single nucleotide polymorphisms (SNP) show high diversity in genomes and they are important markers for map construction and marker assistant selection (MAS). The development of high-throughput sequencing technology can provide the capacity to discover massive numbers of SNP sites in the genome. Polyploid crops have large size genomes along with large scale repetitive sequences, as a result, there are many challenges in developing SNP sites in the polyploidy crops.【Method】300 sweetpotato accessions were collected and sequenced by SLAF-seq technology. The scheme of the experiment was designed based on bio-informatics technology. Taking potato as the reference genome, specific size of DNA were chosen to construct the SLAF-seq library. After high-throughput sequencing, a great amount of sequences were obtained and used to obtain the polymorphism SLAF tags by software alignment, then found the distribution of specific SNP sites. 【Result】 The results coming from the alignment between the sequences of CK Arabidopsis and its reference genome indicated that the construction of SLAF library fitted well to the standard, with its paired-end mapped reads reaching 87.71%, normal digestion ratio reached 93.22%. With the help of sequencing alignment as well as other bio-informatics technologies, a total of 498.14 Mb reads were obtained,the total number of the reads of each sample varied from 441 595 to 2 731 920. Jishu4 had the largest number of reads with 2 731 920 reads. Arabidopsis had the smallest amount of data with 441 595 reads. The quality of Q30 changed from 88.37% to 90.67%, the Q30 of 3043 only reached 87.31%, while the Q30 of Ezi1 reached as high as 91.31%, the average value of Q30 of each sample was above 80%. The GC content of each sample varied from 37.23% to 38.09%; Sushu9 and Zheshu2 had extreme values with Sushu9 having a GC content of 39.80% , while Zheshu2 had 37.10%. The average value of each sample was 37.64% with the average level of GC content low enough to process the sequencing. In the end, 597 094 high quality SLAFs were produced, 260 000 were polymorphisms, accounting for 43.54%. The final results consisted of 795 794 SNP sites obtained based on the analysis of 260 000 polymorphic SLAFs.【Conclusion】A total of 498.14 Mb reads were obtained and produced 597 094 high quality SLAFs, of which 260 000 were polymorphisms. 795 794 SNP sites were developed based on SLAF-seq technology. This paper provided results that SLAF-seq technology can be well applied in developing SNP sites in sweetpotato, much more efficiently than markers like SSR, AFLP, or RAPD. The developed SNP sites can be further used for the analysis of population evolution and developing specific SNP markers.

Key words: sweetpotato, SLAF-seq, SNP sites, high-throughput sequencing, molecular marker

SU Wen-jin, ZHAO Ning, LEI Jian, WANG Lian-jun, CHAI Sha-sha, YANG Xin-sun. SNP Sites Developed by Specific Length Amplification Fragment Sequencing (SLAF-seq) in Sweetpotato[J].Scientia Agricultura Sinica, 2016, 49(1): 27-34.

References

[1] 陆漱韵, 刘庆昌, 李惟基. 甘薯育种学. 北京: 中国农业出版社, 1998: 74.

Lu S Y, Liu Q C, Li W J. Sweetpotato Breeding. Beijing: China Agriculture Press, 1998: 74. (in Chinese)

[2] 夏春丽, 于永利, 张小燕. 甘薯的营养保健作用及开发利用. 食品工程, 2008(3): 28-31.

Xia C L, Yu Y L, Zhang X Y. Nutrition and utilization of sweetpotato. Food Engineering, 2008(3): 28-31. (in Chinese)

[3] Buteler M I, Jarret R L, Labonte D R. Sequence characterization of microsatellites in diploid and polyploid Ipomoea. Theoretical and Applied Genetics, 1999, 99(1/2): 123-132.

[4] Cervantes-Flores J C, Sosinski B, Pecota K V, Mwanga R O M, Catignani G L, Truong V D, Watkins R H, Ulmer M R, Yencho G C. Identification of QTL for dry-matter, starch, and β-carotene content in sweetpotato.Molecular Breeding, 2011, 28(2): 201-216.

[5] Elameen A, Fjellheim S, Larsen A, Rognli O, Sundheim L, Msolla S, Masumba E, Mtunda K, Klemsdal S. Analysis of genetic diversity in a sweetpotato germplasm collection from tanzania as revealed by AFLP.Genetic Resources and Crop Evolution, 2008, 55(3): 397-408.

[6] Liu D G, Zhao N, Zhai H, Yu X X, Jie Q, Wang L J, He S Z, Liu Q C. AFLP fingerprinting and genetic diversity of main sweetpotato varieties in China.Journal of Integrative Agriculture, 2012, 11(9): 1424-1433.

[7] Zhang D P, Cervantes J, Huamán Z, Carey E, Ghislain M. Assessing genetic diversity of sweet potato cultivars from tropical America using AFLP. Genetic Resources and Crop Evolution, 2000, 47(6): 659-665.

[8] Gichuki S T, Berenyi M, Zhang D P, Hermann M, Schmidt J, Glössl J, Burg K. Genetic diversity in sweetpotato in relationship to geographic sources as assessed with RAPD markers. Genetic Resources and Crop Evolution, 2003, 50(4): 429-437.

[9] Jarret R L, Austin D F. Genetic diversity and systematic relationships in sweetpotato and related species as revealed by RAPD analysis.Genetic Resources and Crop Evolution, 1994, 41(3): 165-173.

[10] Zhao N, Yu X X, Jie Q, Li H, Li H, Hu J, Zhai H, He S Z, Liu Q C. A genetic linkage map based on AFLP and SSR markers and mapping of QTL for dry-matter content in sweetpotato. Molecular Breeding, 2013, 32(4): 807-820.

[11] Fajardo D S, Bonte D R L, Jarret R L. Identifying and selecting for genetic diversity in papua new guinea sweetpotato. Genetic Resources and Crop Evolution, 2002, 49(5): 463-470.

[12] 李强, 刘庆昌, 翟红, 马代夫, 王欣, 李雪琴, 王玉萍. 中国甘薯主要亲本遗传多样性的ISSR分析. 作物学报, 2008, 34(6): 972-977.

Li Q, Liu Q C, Zhai H, Ma D F, Wang X, Li X Q, Wang Y P. Genetic diversity in main parents of sweetpotato in China as revealed by ISSR markers. Acta Agronomica Sinica, 2008, 34(6): 972-977. (in Chinese)

[13] Jarret R L, Bowen N. Simple sequence repeats (SSRs) for sweetpotato germplasm characterization. Plant Genetic Resources Newsletter 1994, 100: 9-11.,

[14] Wang Z Y, Li J, Luo Z X, Huang L F, Chen X L, Fang B P, Li Y J, Chen J Y, Zhang X J. Characterization and development of est-derived SSR markers in cultivated sweetpotato. BMC Plant Biology, 2011, 11(1): 1-9.

[15] Yang X S, Su W J, Wang L J, Lei J, Chai S S, Liu Q C. Molecular diversity and genetic structure of 380 sweetpotato accessions as revealed by SSR markers.Journal of Integrative Agriculture, 2015(4): 633-641.

[16] Zhang Y X, Wang L H, Xin H G, Li D H, Ma C X, Ding X, Hong W G, Zhang X R. Construction of a high-density genetic map for sesame based on large scale marker development by specific length amplified fragment (SLAF) sequencing.BMC Plant Biology,2013, 13(1): 1-12.

[17] Sun X W, Liu D Y, Zhang X F, Li W B, Liu H, Hong W G, Jiang C B, Guan N, Ma C X, Zeng H P. SLAF-seq: An efficient method of large-scale de novo snp discovery and genotyping using high-throughput sequencing.Plos One, 2013, 8(3): e58700.

[18] Xun X, Pan S, Cheng S, Zhang B, Mu D, Ni P, Zhang G, Yang S, Li R, Wang J, Orjeda G, Guzman F, Torres M, Lozano R, Ponce O, Martinez D, De la Cruz G, Chakrabarti S K, Patil V U, Skryabin K G, Kuznetsov B B, Ravin N V, Kolganova T V, Beletsky A V, Mardanov A V, Di Genova A, Bolser D M, Martin D M, Li G, Yang Y, Kuang H, Hu Q, Xiong X, Bishop G J, Sagredo B, Mejía N, Zagorski W, Gromadka R, Gawor J, Szczesny P, Huang S, Zhang Z, Liang C, He J, Li Y, He Y, Xu J, Zhang Y, Xie B, Du Y, Qu D, Bonierbale M, Ghislain M, Herrera M R, Giuliano G, Pietrella M, Perrotta G, Facella P, O'Brien K, Feingold S E, Barreiro L E, Massa G A, Diambra L, Whitty B R, Vaillancourt B,Lin H, Massa A N, Geoffroy M, Lundback S, DellaPenna D, Buell C R, Sharma S K, Marshall D F, Waugh R, Bryan G J, Destefanis M, Nagy I, Milbourne D, Thomson S J, Fiers M, Jacobs J M, Nielsen K L, Sønderkær M, Iovene M, Torres G A, Jiang J, Veilleux R E, Bachem C W, de Boer J, Borm T, Kloosterman B, van Eck H, Datema E, Hekkert B T, Goverse A, van Ham R C, Visser R G. Genome sequence and analysis of the tuber crop potato. Nature, 2011, 475(7355): 189-195.

[19] Wei Q Z, Wang Y Z, Qin X D, Zhang Y X, Zhang Z T, Wang J, Li J, Lou Q F, Chen J F. An SNP-based saturated genetic map and QTL analysis of fruit-related traits in cucumber using specific-length amplified fragment (SLAF) sequencing. BMC Genomics, 2014, 15: 1158.

[20] Li B, Tian L, Zhang J Y, Huang L, Han F X, Yan S R, Wang L Z, Zheng H K, Sun J M. Construction of a high-density genetic map based on large-scale markers developed by specific length amplified fragment sequencing (SLAF-seq) and its application to QTL analysis for isoflavone content in Glycine max. BMC Genomics, 2014, 15: 1086.

[21] Zhang Y X, Wang L H, Xin H G, Li D H, Ma C X, Ding X, Hong W G, Zhang X R. Construction of a high-density genetic map for sesame based on large scale marker development by specific length amplified fragment (SLAF) sequencing. BMC Plant Biology, 2013, 13(17): 2636-2646.

[22] Huang S, Ding J, Deng D, Tang W, Sun H, Liu D, Zhang L, Niu X, Zhang X, Meng M, Yu J, Liu J, Han Y, Shi W, Zhang D, Cao S, Wei Z, Cui Y, Xia Y, Zeng H, Bao K, Lin L, Min Y, Zhang H, Miao M, Tang X, Zhu Y, Sui Y, Li G, Sun H, Yue J, Sun J, Liu F, Zhou L, Lei L, Zheng X, Liu M, Huang L, Song J, Xu C, Li J, Ye K, Zhong S, Lu B R, He G, Xiao F, Wang H L, Zheng H, Fei Z, Liu Y. Draft genome of the kiwifruit actinidia chinensis. Nature Communications, 2013, 4: 2640.

[23] 陈士强, 秦树文, 黄泽峰, 戴毅, 张璐璐, 高营营, 高勇, 陈建民. 基于SLAF-seq技术开发长穗偃麦草染色体特异分子标记. 作物学报, 2013, 39(4): 727-734.

Chen S Q, Qin S W, Huang Z F, Dai Y, Zhang L L, Gao Y Y, Gao Y, Chen J M. Development of specific molecular markers for Thinopyrum elongatum chromosome using SLAF-seq technique. Acta Agronomica Sinica, 2013, 39(4): 727-734. (in Chinese)

[24] Chen W, Yao J B, Chu L, Li Y, Guo X M, Zhang Y S. The development of specific SNP markers for chromosome 14 in cotton using next-generation sequencing. Plant Breeding, 2014, 133(2): 256-261.

[25] Zhang J, Zhang J P, Liu W H, Han H M, Lu Y Q, Yang X M, Li X Q, Li L H. Introgression of Agropyron cristatum 6P chromosome segment into common wheat for enhanced thousand-grain weight and spike length. Theoretical and Applied Genetics, 2015, 128(9): 1827-1837.

[26] Zhu Y F, Yin Y F, Yang K Q, Li J H, Sang Y L, Huang L, Fan S. Construction of a high-density genetic map using specific length amplified fragment markers and identification of a quantitative trait locus for anthracnose resistance in walnut (Juglans regia L.). BMC Genomics, 2015, 16(1): 1-13.

[27] Cai C, Cheng F Y, Wu J, Zhong Y, Liu G. The first high-density genetic map construction in Tree Peony (Paeonia Sect. Moutan) using Genotyping by Specific-Locus Amplified Fragment Sequencing. Plos One, 2015, 10(5): e0128584.

[28] Ma J Q, Huang L, Ma C L, Jin J Q, Li C F, Wang R K, Zheng H K, Yao M Z, Chen L. Large-scale SNP discovery and genotyping for constructing a high-density genetic map of tea plant using Specific- Locus Amplified Fragment Sequencing (SLAF-seq). Plos One, 2015, 10(6): e0128798.

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SNP Sites Developed by Specific Length Amplification Fragment Sequencing (SLAF-seq) in Sweetpotato

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