Utility of EST-derived SSR in cultivated peanut (Arachis hypogaea L.) and Arachis wild species - PubMed (original) (raw)

Utility of EST-derived SSR in cultivated peanut (Arachis hypogaea L.) and Arachis wild species

Xuanqiang Liang et al. BMC Plant Biol. 2009.

Abstract

Background: Lack of sufficient molecular markers hinders current genetic research in peanuts (Arachis hypogaea L.). It is necessary to develop more molecular markers for potential use in peanut genetic research. With the development of peanut EST projects, a vast amount of available EST sequence data has been generated. These data offered an opportunity to identify SSR in ESTs by data mining.

Results: In this study, we investigated 24,238 ESTs for the identification and development of SSR markers. In total, 881 SSRs were identified from 780 SSR-containing unique ESTs. On an average, one SSR was found per 7.3 kb of EST sequence with tri-nucleotide motifs (63.9%) being the most abundant followed by di- (32.7%), tetra- (1.7%), hexa- (1.0%) and penta-nucleotide (0.7%) repeat types. The top six motifs included AG/TC (27.7%), AAG/TTC (17.4%), AAT/TTA (11.9%), ACC/TGG (7.72%), ACT/TGA (7.26%) and AT/TA (6.3%). Based on the 780 SSR-containing ESTs, a total of 290 primer pairs were successfully designed and used for validation of the amplification and assessment of the polymorphism among 22 genotypes of cultivated peanuts and 16 accessions of wild species. The results showed that 251 primer pairs yielded amplification products, of which 26 and 221 primer pairs exhibited polymorphism among the cultivated and wild species examined, respectively. Two to four alleles were found in cultivated peanuts, while 3-8 alleles presented in wild species. The apparent broad polymorphism was further confirmed by cloning and sequencing of amplified alleles. Sequence analysis of selected amplified alleles revealed that allelic diversity could be attributed mainly to differences in repeat type and length in the microsatellite regions. In addition, a few single base mutations were observed in the microsatellite flanking regions.

Conclusion: This study gives an insight into the frequency, type and distribution of peanut EST-SSRs and demonstrates successful development of EST-SSR markers in cultivated peanut. These EST-SSR markers could enrich the current resource of molecular markers for the peanut community and would be useful for qualitative and quantitative trait mapping, marker-assisted selection, and genetic diversity studies in cultivated peanut as well as related Arachis species. All of the 251 working primer pairs with names, motifs, repeat types, primer sequences, and alleles tested in cultivated and wild species are listed in Additional File 1.

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Figures

Figure 1

Figure 1

Polyacrylamide gel electrophoresis patterns of microsatellite alleles amplified with the primer EM-31. The bands indicated by the arrows were sequenced. M represents the DNA molecular weight marker, and 1–38 represent PI 393531 (1), PI 390693 (2), Qiongshanhuasheng (3), Liaoningsilihong (4), Dedou (5), Guangliu (6), Sanyuening (7), Yueyou 20 (8), Spancross (9), Tennessee Red (10), Xiaoliuqiu (11), Yangjiangpudizan (12), Xihuagoudo (13), Padou (14), Bo-50 (15), Yingdejidouzai (16), Heyuanbanman (17), Tuosunxiaohuasheng (18), Sunoleic 97R (19), Tifrunner (20), Georgia Green (21), NC940-22 (22), A. villosa (23), A. stenosperma (24), A. correntina (25), A. cardenasii (26), A. magna (27), A. duranensis (28), A. chacoensis (29), A. batizocoi (30), A. helodes (31), A. monticola (32), A. pintoi (33), A. paraguariensis (34), A. pusilla (35), A. rigonii (36), A. appressipila (37), A. glabrata (38).

Figure 2

Figure 2

Alignment of sequences obtained from five SSR bands amplified by EM-31 primers and original SSR-derived EST sequence(EM-31). Primer sequences are indicated by underlined arrows. Repetitive sequences are indicated in dashed box. Point mutations and indel regions are marked by box with solid line.

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References

    1. Krapovickas A, Gregory WC. Taxonomia del genero Arachis (Leguminosae) Bonplandia. 1994;8:1–186.
    1. Halward TM, Stalker HT, Larue EA, Kochert G. Genetic variation detectable with molecular markers among unadapted germplasm resources of cultivated peanut and related wild species. Genome. 1991;34:1013–1020.
    1. Kochert G, Halward T, Branch WD, Simpson CE. RFLP variability in peanut (Arachis hypogaea L.) cultivars and wild species. Theor Appl Genet. 1991;81:565–570. doi: 10.1007/BF00226719. - DOI - PubMed
    1. He GH, Prakash CS. Identification of polymorphic DNA markers in cultivated peanut (Arachis hypogaea L.) Euphytical. 1997;97:143–149. doi: 10.1023/A:1002949813052. - DOI
    1. Subramanian V, Gurtu S, Nageswara Rao RC, Nigam SN. Identification of DNA polymorphism in cultivated groundnut using random amplified polymorphic DNA (RAPD) assay. Genome. 2000;43:656–660. doi: 10.1139/gen-43-4-656. - DOI - PubMed

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