Genome wide identification and characterization of microsatellite markers in black pepper (Piper nigrum): A valuable resource for boosting genomics applications

Autoři: Ratna Kumari aff001;  Dhammaprakash Pandhari Wankhede aff001;  Akansha Bajpai aff001;  Avantika Maurya aff001;  Kartikay Prasad aff001;  Dikshant Gautam aff001;  Parimalan Rangan aff001;  M. Latha aff001;  Joseph John K. aff001;  Suma A. aff001;  Kangila V. Bhat aff001;  Ambika B. Gaikwad aff001
Působiště autorů: ICAR-National Bureau of Plant Genetic Resources, New Delhi, India aff001
Vyšlo v časopise: PLoS ONE 14(12)
Kategorie: Research Article
doi: 10.1371/journal.pone.0226002


Black pepper is one of the most valued and widely used spices in the world and dominates multi-billion dollar global spices trade. India is amongst the major producers, consumers and exporters of black pepper. In spite of its commercial and cultural importance, black pepper has received meagre attention in terms of generation of genomic resources. Availability of markers distributed throughout the genome would facilitate and accelerate genetic studies, QTL identification, genetic enhancement and crop improvement in black pepper. In this perspective, the sequence information from the recently sequenced black pepper (Piper nigrum) genome has been used for identification and characterisation of Simple Sequence Repeats (SSRs). Total 69,126 SSRs were identified from assembled genomic sequence of P. nigrum. The SSR frequency was 158 per MB making it, one SSR for every 6.3 kb in the assembled genome. Among the different types of microsatellite repeat motifs, dinucleotides were the most abundant (48.6%), followed by trinucleotide (23.7%) and compound repeats (20.62%). A set of 85 SSRs were used for validation, of which 74 produced amplification products of expected size. Genetic diversity of 30 black pepper accessions using 50 SSRs revealed four distinct clusters. Further, the cross species transferability of the SSRs was checked in nine other Piper species. Out of 50 SSRs used, 19 and 31 SSRs were amplified in nine and seven species, respectively. Thus the identified SSRs may have application in other species of the genus Piper where genome sequence is not available yet. Present study reports the first NGS based genomic SSRs in black pepper and thus constitute a valuable resource for a whole fleet of applications in genetics and plant breeding studies such as genetic map construction, QTL identification, map-based gene cloning, marker-assisted selection and evolutionary studies in Piper nigrum and related species.

Klíčová slova:

Comparative genomics – Crop genetics – Evolutionary genetics – Gene mapping – Genomic libraries – India – Microsatellite loci – Sequence motif analysis


1. Nair KP (2011) Agronomy and Economy of Black Pepper and Cardamom: The" king" and" queen" of Spices. Elsevier; eBook ISBN: 9780123918772.

2. Ravindran PN, Kallupurackal JA (2012) Handbook of Herbs and Spices (Second Edition) Wood head Publishing Series in Food Science, Technology and Nutrition.

3. Srinivasan K (2007) Black pepper and its pungent principle—Piperine: A review of diverse physiological effects. Crit Rev Food Sci Nutr 47: 735–748. doi: 10.1080/10408390601062054 17987447

4. Tsai IL, Lee FP, Wu CC, Duh CY, Ishikawa T, et al. (2005) New cytotoxic cyclobutanoid amides, anew furanoid lignan and anti-platelet aggregation constituents from Piper arborescens, Planta Medica 71: 535–542. doi: 10.1055/s-2005-864155 15971125

5. Selvendiran K, Padmavathi R, Magesh V, Sakthisekaran D, (2005) Preliminary study on inhibition of genotoxicity by Piperine in mice. Fitoterapia 76: 296–300. doi: 10.1016/j.fitote.2005.03.016 15890459

6. Platel K, Srinivasan K (2004) Digestive stimulant action of spices: A myth or reality? Ind J Med Res 119: 167–179.

7. YES BANK and All India Spices Exporters Forum (2018) Indian Spices Industry: Opportunities in Domestic & Global Markets.

8. Ravindran PN. BLACK PEPPER (Piper nigrum), Indian Institute of Spices Research, Kozhikode, CRC Press; 2000

9. Raghavan R, Elumalai S, Babu KN, Hittalmani S (2010) Molecular Characterization of Black Pepper (Piper Nigrum) Using RAPD and SSR Markers. Biosci Biotech Res Asia 7:2

10. Joy N, Prasanth VP, Soniya EV (2011) Microsatellite based analysis of genetic diversity of popular black pepper genotypes in South India. Genetica 139:1033–43. doi: 10.1007/s10709-011-9605-x 21874534

11. Jiang Y and Liu JP (2011). Analysis of genetic diversity of Piper spp. in Hainan Island (China) using inter-simple sequence repeat ISSR markers. Afr J Biotechnol 10: 14731–14737.

12. Jiang Y, Liu JP (2011) Evaluation of genetic diversity in Piper spp using RAPD and SRAP markers. Genet Mol Res 10: 2934–2943. doi: 10.4238/2011.November.29.4 22179965

13. Wu BD, Fan R, Hu LS, Wu HS, Hao CY (2016) Genetic diversity in the germplasm of black pepper determined by EST-SSR markers. Genet Mol Res 15:1.

14. Kole C, Muthamilarasan M, Henry R, Edwards D, Sharma R, et al. (2015) Application of genomics-assisted breeding for generation of climate resilient crops: progress and prospects Front Plant Sci 6: 563. doi: 10.3389/fpls.2015.00563 26322050

15. Ren Z, Gao J, Li LG, Cai X, Huang W, Chao D, et al. (2005) A rice quantitative trait locus for salt tolerance encodes a sodium transporter. Nat Genet 37: 1141–1146. doi: 10.1038/ng1643 16155566

16. Reddy UK, Abburi L, Abburi VL, Saminathan T, Cantrell R, et al. (2015) A genome-wide scan of selective sweeps and association mapping of fruit traits using microsatellite markers in watermelon. J Hered 106:166–76. doi: 10.1093/jhered/esu077 25425675

17. Lambel S, Lanini B, Vivoda E, Fauve J, Wechter et al. (2014) A major QTL associated with Fusarium oxysporum race 1resistance identified in genetic populations derived from closely related watermelon lines using selective genotyping and genotyping-by sequencing for SNP discovery. Theor Appl Genet 127:2105–15. doi: 10.1007/s00122-014-2363-2 25104326

18. Ren Y, Jiao D, Gong GY, Zhang HY, Guo SG, et al. (2015) Genetic analysis and chromosome mapping of resistance to Fusarium oxysporum f. sp niveum (FON) race 1 and race 2 in watermelon (Citrullus lanatus L.). Mol Breeding 35:183.

19. Yang L, Koo DH, Li D, Zhang T, Jiang J, et al. (2014) Next-generation sequencing, FISH mapping and synteny-based modeling reveal mechanisms of decreasing dysploidy in Cucumis. Plant J 77:16–30. doi: 10.1111/tpj.12355 24127692

20. Varshney RK, Graner A, Sorrells ME (2005) Genic microsatellite markers in plants: features and applications. Trends Biotechnol 23: 48–55. doi: 10.1016/j.tibtech.2004.11.005 15629858

21. Tautz D, Renz M (1984) Simple sequences are ubiquitous repetitive components of eukaryotic genomes. Nucleic Acids Res. 12: 4127–4138. doi: 10.1093/nar/12.10.4127 6328411

22. Menezes IC, Cidade FW, Souza EAP, Sampaio EIC (2009) Isolation and characterization of microsatellite loci in the black pepper, Piper nigrum L. (Piperaceae) Conservation Genet Resour 1:209–212.

23. Gordo SM, Pinheiro DG, Moreira EC, Rodrigues SM, Poltronieri MC, et al. (2012) High-throughput sequencing of black pepper root transcriptome. BMC Plant Biol. 12:168. doi: 10.1186/1471-2229-12-168 22984782

24. Hu L, Hao C, Fan R, Wu B, Tan L, Wu H (2015) De Novo Assembly and Characterization of Fruit Transcriptome in Black Pepper (Piper nigrum). PLoS One. 10:e0129822. doi: 10.1371/journal.pone.0129822 26121657

25. Mardis ER (2008): The impact of next-generation sequencing technology on genetics. Trends Genet 24: 133–141. doi: 10.1016/j.tig.2007.12.007 18262675

26. Yang T, Bao SY, Ford R, Jia TJ, Guan JP, et al. (2012) High-throughput novel microsatellite marker of faba bean via next generation sequencing. BMC Genomics 13:602. doi: 10.1186/1471-2164-13-602 23137291

27. Zhu H, Song P, Koo DH, Guo L, Li Y, Sun S, et al. (2016) Genome wide characterization of simple sequence repeats in watermelon genome and their application in comparative mapping and genetic diversity analysis. BMC Genomics 17:557. doi: 10.1186/s12864-016-2870-4 27495254

28. Wang Q, Fang L, Chen J, Hu Y, Si Z, et al. (2015) Genome-wide mining, characterization, and development of microsatellite markers in Gossypium species. Sci Rep 5:10638. doi: 10.1038/srep10638 26030481

29. Gimode D, Odeny DA, de Villiers EP, Wanyonyi S, Dida MM, et al. (2016) Identification of SNP and SSR Markers in Finger Millet Using Next Generation Sequencing Technologies. PLoS One. 11:e0159437. doi: 10.1371/journal.pone.0159437 27454301

30. Pandey G, Misra G, Kumari K, Gupta S, Parida SK, et al. (2013) Genome-wide development and use of microsatellite markers for large-scale genotyping applications in foxtail millet [Setaria italica (L.)]. DNA Research 20: 197–207. doi: 10.1093/dnares/dst002 23382459

31. Zhao C, Qiu J, Agarwal G, Wang J, Ren X, et al. (2017) Genome- wide discovery of microsatellite markers from diploid progenitor species, Arachis duranensis and A. Ipaensis, and their application in cultivated peanut (a. Hypogaea). Front Plant Sci 8: 1209. doi: 10.3389/fpls.2017.01209 28769940

32. Bastías A, Correa F, Rojas P, Almada R, Muñoz C, Sagredo B (2016) Identification and Characterization of Microsatellite Loci in Maqui (Aristotelia chilensis [Molina] Stunz) Using Next-Generation Sequencing (NGS). PLoS One.11: e0159825. doi: 10.1371/journal.pone.0159825 27459734

33. Barbara T, Palma-Silva CL, Paggi GM, Bered F, Fay MF, Lexer C (2007). Cross species transfer of nuclear microsatellite markers: potential and limitations. Mol Eco 16:3759–67.

34. Xiao Y, Xia W, Ma J, Mason AS, Fan H, et al. (2016) Genome-wide identification and transferability of microsatellite markers between Palmae species. Front in Plant Sci. 7: 1578.

35. Shiferaw E (2013) Development and cross-species amplification of grass pea EST-derived markers. Afri Crop Sci Jour. 21:153–60.

36. Wang H, Walla JA, Zhong S, Huang D, Dai W (2012) Development and cross-species/ genera transferability of microsatellite markers discovered using 454 genome sequencing in chokecherry (Prunus virginiana L.). Plant cell reports 1:31.

37. Datta S, Mahfooz S, Singh P, Choudhary AK, Singh F, Kumar S (2010) Cross-genera amplification of informative microsatellite markers from common bean and lentil for the assessment of genetic diversity in pigeonpea. Physio Mol Biol of Plants16: 123–34.

38. Guo W, Wang W, Zhou B, Zhang T (2006) Cross-species transferability of G. arboreum-derived EST-SSRs in the diploid species of Gossypium. Theor Appl Genet 112:1573–81. doi: 10.1007/s00122-006-0261-y 16596396

39. Ramasamy Y, Ghosh M, Sumathi R, Gurumurthi K(2005) Cross-species amplification of eucalyptus SSR markers in Casuarinaceae. Acta Bot Croat 64:115–120.

40. Pandian A, Ford R, Taylor PW (2000) Transferability of sequence tagged microsatellite site (STMS) primers across four major pulses. Plant Mol Biol Reporter 1:18.

41. Saghai-Maroof MA, Soliman KM, Jorgensen RA, Allard RW (1984) Ribosomal DNA spacer-length polymorphism in barley: Mendelian inheritance, chromosomal location, and population dynamics. PNAS 81:8014–8019. doi: 10.1073/pnas.81.24.8014 6096873

42. Beier S, Thiel T, Münch T, Scholz U, Mascher M (2017) MISA-web: a web server for microsatellite prediction. Bioinformatics 33: 2583–2585 doi: 10.1093/bioinformatics/btx198 28398459

43. Rozen S, Skaletsky H (2000) Primer3 on theWWWfor general users and for biologist programmers. Methods in molecular biology 132: 365–386. doi: 10.1385/1-59259-192-2:365 10547847

44. Rohlf FJ (2000) NTSYS-pc Numerical taxonomy and multivariate system, version 2.1. Applied Biostatics Inc., New York

45. La Rota M, Kantety RV, Yu JK, Sorrells ME (2005) Nonrandom distribution and frequencies of genomic and EST-derived microsatellite markers in rice, wheat, and barley. BMC Genomics 6:23. doi: 10.1186/1471-2164-6-23 15720707

46. Chabane K, Ablett GA, Cordeiro GM, Valkoun J, Henry RJ (2005) EST versus genomic derived microsatellite markers for genotyping wild and cultivated barley. Genet Res Crop Evol 52:903–909

47. Pinto LR, Oliveira KM, Marconi T, Garcia AAF, Ulian EC, De Souza AP (2006) Characterization of novel sugarcane expressed sequence tag microsatellites and their comparison with genomic SSRs. Plant Breed 125: 378–384

48. Wang YW, Samuels TD, Wu YQ (2011) Development of 1,030 genomic SSR markers in switchgrass. Theor Appl Genet 122: 677–86. doi: 10.1007/s00122-010-1477-4 20978736

49. Warnke SE, Barker RE, Jung G, Sim SC, Mian MAR, et al. (2004) Genetic linkage mapping of an annual x perennial ryegrass population. Theor Appl Genet 109: 294–304. doi: 10.1007/s00122-004-1647-3 15071730

50. Kalia RK, Rai MK, Kalia S, Singh R, Dhawan AK (2011) Microsatellite markers: an overview of the recent progress in plants. Euphytica, 177: 309–334.

51. Tangphatsornruang S, Somta P, Uthaipaisanwong P, Chanprasert J, Sangsrakru D, et al. (2009) Characterization of microsatellites and gene contents from genome shotgun sequences of mungbean (Vigna radiata (L.) Wilczek). BMC Plant Biology 9: 137. doi: 10.1186/1471-2229-9-137 19930676

52. Zhu H, Senalik D, McCown BH, Zeldin EL, Speers J, et al. (2012) Mining and validation of pyrosequenced simple sequence repeats (SSRs) from American cranberry (Vaccinium macrocarpon Ait.). Theoretical and applied genetics 124: 87–96. doi: 10.1007/s00122-011-1689-2 21904845

53. Dutta S, Kumawat G, Singh BP, Gupta DK, Singh S, et al. (2011) Development of genic-SSR markers by deep transcriptome sequencing in pigeonpea [Cajanus cajan (L.) Millspaugh]. BMC Plant Biology 11: 17. doi: 10.1186/1471-2229-11-17 21251263

54. Anupama K, Anu Cyriac, Saji KV, Nirmal Babu K (2015) Microsatellite marker based cross species amplification and genetic diversity analysis in the genus Piper. Inter Jour of Adv Resear 3: 184–91.

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