Development and Characterization of EST-SSR Molecular Markers in Larix kaempferi

LIU Chao; ZHANG Li-peng; WANG Chun-guo; SONG Wen-qin; CHEN Cheng-bin

Volume 26 Issue S1

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Development and Characterization of EST-SSR Molecular Markers in Larix kaempferi

1.
College of Life Sciences, Nankai University, Tianjin 300071, China

Received Date: 2013-08-23

Abstract

Based on RNA sequences information of Larix kaempferi, 1 788 EST-SSR loci were found by SSRIT, with an average length of 17.5 bp. The most abundant SSRs within Larix were hexanucleotide and trinucleotide, followed by pentanucleotide, dinucleotide, and tetranucleotide. At the meantime, 656 kinds of repeat primitives were detected. In this study, a total of 500 primers were designed by "Primer Premier 5.0" and 102 pairs were synthesized. These SSR primers were used to assess the amplification efficiency of Taxaceae, Cupressaceae, and Pinaceae. The results showed that 95% and 50% of all primers could be amplified successfully in Taxaceae and Cupressaceae. And in Pinaceae, the transferability of Larix SSR in Larix, Pseudotsuga, Abies, Cedrus, Picea and Pinus was 99.01%, 67.4%., 67.4%, 63.8%, 70.2%, and 75.7% respectively. Seven different varieties of Larix genetic relationship were analyzed by using 101 pairs of SSR primers. 79 polymorphism loci were obtained, and Larix was divided into four groups at the genetic similarity coefficient of 0.69.
- Larix kaempferi(Lamb.)Carr.,
- transcriptome sequencing,
- genic-SSR,
- transferability

References

[1]	Kalia, R K,Rai M K,Kalia S, et al. Microsatellite markers: an overview of the recent progress in plants[J]. Euphytica,2011,177(3):309-334
[2]	李小白,向林,罗洁,等.转录组测序(RNA-seq)策略及其数据在分子标记开发上的应用[J]. 中国细胞生物学学报,2013, 35(5):1-7
[3]	Luro F L, Costantino G, Terol J, et al. Transferability of the EST-SSRs developed on Nules clementine (Citrus clementina Hort ex Tan) to other Citrus species and their effectiveness for genetic mapping[J]. BMC Genomics, 2008, 9: 287-291
[4]	Zeng S, Xiao G, Guo J, et al. Development of a EST dataset and characterization of EST-SSRs in a traditional Chinese medicinal plant, Epimedium sagittatum (Sieb.Et Zucc.) Maxim[J]. BMC Genomics, 2010, 11: 94
[5]	Zhang H, Wei L, Miao H, et al. Development and validation of genic-SSR markers in sesame by RNA-seq[J]. BMC Genomics, 2012, 13: 316
[6]	Sakaguchi S, Uchiyama K, Ueno S, et al. Isolation and characterization of 52 polymorphic EST-SSR markers for Callitris columellaris (Cupressaceae)[J]. Am J Bot, 2011, 98(12): e363-e368
[7]	Ueno S, Moriguchi Y, Uchiyama K, et al. A second generation framework for the analysis of microsatellites in expressed sequence tags and the development of EST-SSR markers for a conifer, Cryptomeria japonica[J]. BMC Genomics, 2012, 13: 136
[8]	Li D, Deng Z, Qin B, et al. Denovo assembly and characterization of bark transcriptome using Illumina sequencing and development of EST-SSR markers in rubber tree (Hevea brasiliensis Muell. Arg.)[J]. BMC Genomics, 2012, 13(1): 192
[9]	Zhang J, Liang S, Duan J, et al. De novo assembly and characterisation of the transcriptome during seed development, and generation of genic-SSR markers in peanut (Arachis hypogaea L.)[J]. BMC Genomics, 2012, 13: 90
[10]	Liang X, Chen X, Hong Y, et al. Utility of EST-derived SSR in cultivated peanut (Arachis hypogaea L.) and Arachis wild species[J]. BMC Plant Biol, 2009, 9: 35
[11]	Schafleitner R, Tincopa L R, Palomino O, et al. A sweetpotato gene index established by de novo assembly of pyrosequencing and Sanger sequences and mining for gene-based microsatellite markers[J]. BMC Genomics, 2010, 11(1): 604
[12]	Dutta S, Kumawat G, Singh B P, et al. Development of genic-SSR markers by deep transcriptome sequencing in pigeonpea [J]. BMC Plant Biol, 2011, 11: 17
[13]	Zhang J, Zhang S, Li S, et al. A genome-wide survey of microRNA truncation and 3' nucleotide addition events in larch (Larix leptolepis). Planta, 2013, 237(4): 1047-1056
[14]	Isoda K,Watanabe A.Isolation and characterization of microsatellite loci from Larix keampferi[J]. Molecular Ecology Notes,2006,6(3) : 664-666
[15]	杨秀艳,孙晓梅,张守攻,等. 日本落叶松 EST-SSR 标记开发及二代优树遗传多样性分析[J]. 林业科学,2012, 47(11): 52-58
[16]	贯春雨,张含国,张磊,等. 基于松科树种 EST 序列的落叶松SSR引物开发[J]. 东北林业大学学报,2011, 39(1):20-23
[17]	Temnykh S, DeClerck G,Lukashova A, et al. Computational and experimental analysis of microsatellites in rice (Oryza sativa L.): frequency, length variation, transposon associations, and genetic marker potential[J]. Genome Research,2001, 11(8):1441-1452
[18]	任亮,朱宝芹,张轶博, 等. 利用软件Primer Premier 5.0进行PCR引物设计的研究[J]. 锦州医学院学报,2004, 25(6):43-46
[19]	翟中会,陈希南,王娟. 利用Primer Premier5.0 进行引物设计[J]. 西北医学教育,2008, 16(4):695-698
[20]	Zhang G, Guo G, Hu X, et al. Deep RNA sequencing at single base-pair resolution reveals high complexity of the rice transcriptome[J]. Genome Res, 2010,20(5): 646-654
[21]	Li X, Sun H, Pei J, et al. De novo sequencing and comparative analysis of the blueberry transcriptome to discover putative genes related to antioxidants. Gene, 2012, 511 (1): 54-61
[22]	Qiu Q, Ma T, Hu Q, et al. Genomescale transcriptome analysis of the desert poplar, Populus euphratica[J]. Tree Physiol,2011, 31(4): 452-461
[23]	Xu D L, Long H, Liang J J, et al. De novo assembly and characterization of the root transcriptome of Aegilops variabilis during an interaction with the cereal cyst nematode[J]. BMC Genomics, 2012,13:133
[24]	Zhang J, Liang S, Duan J, et al. Denovo assembly and Characterisation of the Transcriptome during seed development, and generation of genic-SSR markers in Peanut (Arachis hypogaea L.)[J]. BMC Genomics,2012,13: 90
[25]	Bérubé Y,Zhuang J,Rungis D, et al. Characterization of EST-SSRs in loblolly pine and spruce[J]. Tree Genetics & Genomes,2007, 3(3):251-259
[26]	王艳敏,魏志刚,杨传平.白桦 EST-SSR 信息分析与标记的开发[J]. 林业科学,2008, 44(2):78-84
[27]	Nicot N,Chiquet V, Gandon B, et al. Study of simple sequence repeat (SSR) markers from wheat expressed sequence tags (ESTs) [J]. Theor. Appl. Genet, 2004, 109(4):800-805
[28]	Yu F,Wang B H,Feng S P, et al. Development, characterization, and cross-species/genera transferability of SSR markers for rubber tree (Hevea brasiliensis)[J]. Plant Cell Reports,2011, 30(3):335-344
[29]	Poncet V,Rondeau M,Tranchant C, et al. SSR mining in coffee tree EST databases: potential use of EST-SSRs as markers for the Coffea genus[J]. Molecular Genetics and Genomics,2006,276(5):436-449
[30]	孔秋生. 基于公共序列数据库的Cucumis属EST-SSR标记的鉴定开发和利用[D]. 武汉: 华中农业大学,2006
[31]	Gupta P,Rustgi S,Sharma S, et al. Transferable EST-SSR markers for the study of polymorphism and genetic diversity in bread wheat[J]. Molecular Genetics and Genomics,2003,270(4): 315-323
[32]	Tang S,Okashah R A, M M. Cordonnier-Pratt, et al. EST and EST-SSR marker resources for Iris. BMC [J].Plant Biology,2009, 9(1):72
[33]	Rajendrakumar P,Biswal A K,Balachandran S M, et al. Simple sequence repeats in organellar genomes of rice: frequency and distribution in genic and intergenic regions[J]. Bioinformatics,2007,23(1):1-4
[34]	Areshchenkova T,Ganal M. Comparative analysis of polymorphism and chromosomal location of tomato microsatellite markers isolated from different sources[J]. Theor. Appl. Genet,2002,104(2-3):229-235
[35]	李红英. 鹅掌楸EST-SSR在木兰科属间通用性及系统学探讨[D]. 江苏南京:南京林业大学, 2008
[36]	廖娇,黄春辉,辜青青,等. 猕猴桃 EST-SSR 引物筛选及通用性分析[J]. 果树学报,2012, 28(6): 1111-1116
[37]	Wen M, Wang H, Xia Z, et al. Development of EST-SSR and genomic-SSR markers to assess genetic diversity in Jatropha Curcas L. BMC Res Notes, 2010, 3(1): 42
[38]	Zhou Y, Gao F, Liu R, et al. De novo sequencing and analysis of root transcriptome using 454 pyrosequencing to discover putative genes associated with drought tolerance in Ammopiptanthus mongolicus. BMC Genomics, 2012, 13(1): 266
[39]	Zhang Pingzhi,Dreisigacker S,Melchinger A E, et al. Quantifying novel sequence variation and selective advantage in synthetic hexaploid wheat and their backcross-derived lines using SSR markers[J].Molecular Breeding,2005, 15(1):1-10
[40]	Eujayl I, Sorrells M E, Wolters P, et al. Isolation of EST-derived microsatellite marker for genotying the A and B genomes of wheat[J]. Theor. Appl. Genet, 2002, 104: 399-407
[41]	Scott K D, Eggler P, Seaton G, et al. Analysis of SSRs derived from grape ESTs[J]. Theor Appl Genet,2000, 100(5): 723-726

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通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

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Development and Characterization of EST-SSR Molecular Markers in Larix kaempferi

1. College of Life Sciences, Nankai University, Tianjin 300071, China

Received Date: 2013-08-23

Abstract: Based on RNA sequences information of Larix kaempferi, 1 788 EST-SSR loci were found by SSRIT, with an average length of 17.5 bp. The most abundant SSRs within Larix were hexanucleotide and trinucleotide, followed by pentanucleotide, dinucleotide, and tetranucleotide. At the meantime, 656 kinds of repeat primitives were detected. In this study, a total of 500 primers were designed by "Primer Premier 5.0" and 102 pairs were synthesized. These SSR primers were used to assess the amplification efficiency of Taxaceae, Cupressaceae, and Pinaceae. The results showed that 95% and 50% of all primers could be amplified successfully in Taxaceae and Cupressaceae. And in Pinaceae, the transferability of Larix SSR in Larix, Pseudotsuga, Abies, Cedrus, Picea and Pinus was 99.01%, 67.4%., 67.4%, 63.8%, 70.2%, and 75.7% respectively. Seven different varieties of Larix genetic relationship were analyzed by using 101 pairs of SSR primers. 79 polymorphism loci were obtained, and Larix was divided into four groups at the genetic similarity coefficient of 0.69.

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Reference (41)

[1]	Kalia, R K,Rai M K,Kalia S, et al. Microsatellite markers: an overview of the recent progress in plants[J]. Euphytica,2011,177(3):309-334
[2]	李小白,向林,罗洁,等.转录组测序(RNA-seq)策略及其数据在分子标记开发上的应用[J]. 中国细胞生物学学报,2013, 35(5):1-7
[3]	Luro F L, Costantino G, Terol J, et al. Transferability of the EST-SSRs developed on Nules clementine (Citrus clementina Hort ex Tan) to other Citrus species and their effectiveness for genetic mapping[J]. BMC Genomics, 2008, 9: 287-291
[4]	Zeng S, Xiao G, Guo J, et al. Development of a EST dataset and characterization of EST-SSRs in a traditional Chinese medicinal plant, Epimedium sagittatum (Sieb.Et Zucc.) Maxim[J]. BMC Genomics, 2010, 11: 94
[5]	Zhang H, Wei L, Miao H, et al. Development and validation of genic-SSR markers in sesame by RNA-seq[J]. BMC Genomics, 2012, 13: 316
[6]	Sakaguchi S, Uchiyama K, Ueno S, et al. Isolation and characterization of 52 polymorphic EST-SSR markers for Callitris columellaris (Cupressaceae)[J]. Am J Bot, 2011, 98(12): e363-e368
[7]	Ueno S, Moriguchi Y, Uchiyama K, et al. A second generation framework for the analysis of microsatellites in expressed sequence tags and the development of EST-SSR markers for a conifer, Cryptomeria japonica[J]. BMC Genomics, 2012, 13: 136
[8]	Li D, Deng Z, Qin B, et al. Denovo assembly and characterization of bark transcriptome using Illumina sequencing and development of EST-SSR markers in rubber tree (Hevea brasiliensis Muell. Arg.)[J]. BMC Genomics, 2012, 13(1): 192
[9]	Zhang J, Liang S, Duan J, et al. De novo assembly and characterisation of the transcriptome during seed development, and generation of genic-SSR markers in peanut (Arachis hypogaea L.)[J]. BMC Genomics, 2012, 13: 90
[10]	Liang X, Chen X, Hong Y, et al. Utility of EST-derived SSR in cultivated peanut (Arachis hypogaea L.) and Arachis wild species[J]. BMC Plant Biol, 2009, 9: 35
[11]	Schafleitner R, Tincopa L R, Palomino O, et al. A sweetpotato gene index established by de novo assembly of pyrosequencing and Sanger sequences and mining for gene-based microsatellite markers[J]. BMC Genomics, 2010, 11(1): 604
[12]	Dutta S, Kumawat G, Singh B P, et al. Development of genic-SSR markers by deep transcriptome sequencing in pigeonpea [J]. BMC Plant Biol, 2011, 11: 17
[13]	Zhang J, Zhang S, Li S, et al. A genome-wide survey of microRNA truncation and 3' nucleotide addition events in larch (Larix leptolepis). Planta, 2013, 237(4): 1047-1056
[14]	Isoda K,Watanabe A.Isolation and characterization of microsatellite loci from Larix keampferi[J]. Molecular Ecology Notes,2006,6(3) : 664-666
[15]	杨秀艳,孙晓梅,张守攻,等. 日本落叶松 EST-SSR 标记开发及二代优树遗传多样性分析[J]. 林业科学,2012, 47(11): 52-58
[16]	贯春雨,张含国,张磊,等. 基于松科树种 EST 序列的落叶松SSR引物开发[J]. 东北林业大学学报,2011, 39(1):20-23
[17]	Temnykh S, DeClerck G,Lukashova A, et al. Computational and experimental analysis of microsatellites in rice (Oryza sativa L.): frequency, length variation, transposon associations, and genetic marker potential[J]. Genome Research,2001, 11(8):1441-1452
[18]	任亮,朱宝芹,张轶博, 等. 利用软件Primer Premier 5.0进行PCR引物设计的研究[J]. 锦州医学院学报,2004, 25(6):43-46
[19]	翟中会,陈希南,王娟. 利用Primer Premier5.0 进行引物设计[J]. 西北医学教育,2008, 16(4):695-698
[20]	Zhang G, Guo G, Hu X, et al. Deep RNA sequencing at single base-pair resolution reveals high complexity of the rice transcriptome[J]. Genome Res, 2010,20(5): 646-654
[21]	Li X, Sun H, Pei J, et al. De novo sequencing and comparative analysis of the blueberry transcriptome to discover putative genes related to antioxidants. Gene, 2012, 511 (1): 54-61
[22]	Qiu Q, Ma T, Hu Q, et al. Genomescale transcriptome analysis of the desert poplar, Populus euphratica[J]. Tree Physiol,2011, 31(4): 452-461
[23]	Xu D L, Long H, Liang J J, et al. De novo assembly and characterization of the root transcriptome of Aegilops variabilis during an interaction with the cereal cyst nematode[J]. BMC Genomics, 2012,13:133
[24]	Zhang J, Liang S, Duan J, et al. Denovo assembly and Characterisation of the Transcriptome during seed development, and generation of genic-SSR markers in Peanut (Arachis hypogaea L.)[J]. BMC Genomics,2012,13: 90
[25]	Bérubé Y,Zhuang J,Rungis D, et al. Characterization of EST-SSRs in loblolly pine and spruce[J]. Tree Genetics & Genomes,2007, 3(3):251-259
[26]	王艳敏,魏志刚,杨传平.白桦 EST-SSR 信息分析与标记的开发[J]. 林业科学,2008, 44(2):78-84
[27]	Nicot N,Chiquet V, Gandon B, et al. Study of simple sequence repeat (SSR) markers from wheat expressed sequence tags (ESTs) [J]. Theor. Appl. Genet, 2004, 109(4):800-805
[28]	Yu F,Wang B H,Feng S P, et al. Development, characterization, and cross-species/genera transferability of SSR markers for rubber tree (Hevea brasiliensis)[J]. Plant Cell Reports,2011, 30(3):335-344
[29]	Poncet V,Rondeau M,Tranchant C, et al. SSR mining in coffee tree EST databases: potential use of EST-SSRs as markers for the Coffea genus[J]. Molecular Genetics and Genomics,2006,276(5):436-449
[30]	孔秋生. 基于公共序列数据库的Cucumis属EST-SSR标记的鉴定开发和利用[D]. 武汉: 华中农业大学,2006
[31]	Gupta P,Rustgi S,Sharma S, et al. Transferable EST-SSR markers for the study of polymorphism and genetic diversity in bread wheat[J]. Molecular Genetics and Genomics,2003,270(4): 315-323
[32]	Tang S,Okashah R A, M M. Cordonnier-Pratt, et al. EST and EST-SSR marker resources for Iris. BMC [J].Plant Biology,2009, 9(1):72
[33]	Rajendrakumar P,Biswal A K,Balachandran S M, et al. Simple sequence repeats in organellar genomes of rice: frequency and distribution in genic and intergenic regions[J]. Bioinformatics,2007,23(1):1-4
[34]	Areshchenkova T,Ganal M. Comparative analysis of polymorphism and chromosomal location of tomato microsatellite markers isolated from different sources[J]. Theor. Appl. Genet,2002,104(2-3):229-235
[35]	李红英. 鹅掌楸EST-SSR在木兰科属间通用性及系统学探讨[D]. 江苏南京:南京林业大学, 2008
[36]	廖娇,黄春辉,辜青青,等. 猕猴桃 EST-SSR 引物筛选及通用性分析[J]. 果树学报,2012, 28(6): 1111-1116
[37]	Wen M, Wang H, Xia Z, et al. Development of EST-SSR and genomic-SSR markers to assess genetic diversity in Jatropha Curcas L. BMC Res Notes, 2010, 3(1): 42
[38]	Zhou Y, Gao F, Liu R, et al. De novo sequencing and analysis of root transcriptome using 454 pyrosequencing to discover putative genes associated with drought tolerance in Ammopiptanthus mongolicus. BMC Genomics, 2012, 13(1): 266
[39]	Zhang Pingzhi,Dreisigacker S,Melchinger A E, et al. Quantifying novel sequence variation and selective advantage in synthetic hexaploid wheat and their backcross-derived lines using SSR markers[J].Molecular Breeding,2005, 15(1):1-10
[40]	Eujayl I, Sorrells M E, Wolters P, et al. Isolation of EST-derived microsatellite marker for genotying the A and B genomes of wheat[J]. Theor. Appl. Genet, 2002, 104: 399-407
[41]	Scott K D, Eggler P, Seaton G, et al. Analysis of SSRs derived from grape ESTs[J]. Theor Appl Genet,2000, 100(5): 723-726

Development and Characterization of EST-SSR Molecular Markers in Larix kaempferi

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通讯作者: 陈斌, bchen63@163.com

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Development and Characterization of EST-SSR Molecular Markers in Larix kaempferi

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