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Title: LEGUME MARKER DEVELOPMENT THROUGH SEQUENCE ANALYSIS USING PRIMERS DERIVED FROM SOYBEAN UNIGENES

Author
item CHOI, IK-YOUNG - KOREA
item Hyten, David
item MATUKUMALLI, LAKSHMI - GEORGE MASON UNIVERSITY
item YOON, MUN-SUP - KOREA
item Cregan, Perry

Submitted to: Mid Atlantic Plant Molecular Biology Society Conference
Publication Type: Abstract Only
Publication Acceptance Date: 11/30/2005
Publication Date: 11/30/2005
Citation: Choi, I., Hyten, D.L., Matukumalli, L.K., Yoon, M., Cregan, P.B. 2005. Legume marker development through sequence analysis using primers derived from soybean unigenes [abstract]. Mid Atlantic Plant Molecular Biology Society Conference .

Interpretive Summary:

Technical Abstract: PCR primers designed to 3 prime-end sequence of 1204 soybean (Glycine max) unigenes with one primers positioned in the 3 prime UTR successfully amplified a single product from soybean genomic DNA from which high quality DNA sequence data were obtained. Also, 135 primers derived from soybean coding sequence successfully amplified a single amplicon that yielded high quality sequence data. We tested these primers on a number of Legume species to determine the success with which primers to soybean genic sequence could serve as cross-Legume sequence tagged sites (STS). The 1204 + 135 STS were used to attempt amplification in other Legume species including common bean (Phaseolus vulgaris), cowpea (Vigna ungiculata), chickpea (Cicer arietinum), pea (Pisum sativum), peanut (Arachis hypogea), barrel medic (Medicago truncatula) and Lotus japonicus. Primers to the STS were used to amplify two genotypes (mapping parents) of each species to determine the level of sequence polymorphism and to discover mappable single nucleotide polymorphisms (SNPs). Sequencable products derived from primers designed to the 3 prime-end soybean unigene sequence were obtained for 15.6, 14.0, 6.2, 5.6, 2.7, 2.7 and 2.1% of the 1204 primer sets in the aforementioned Legume species, respectively. Sequencable products were obtained from 35.6, 28.1, 25.2, 20.7, 20.0, 22.2, and 25.9% of the 135 primer sets derived from coding sequence via amplification of genomic DNA of P. vulgaris, V. ungiculata, C. arietinum, M. truncatula, P. sativum, A. hypogea and L. japonicus, respectively. Thirty-nine primer sets were discovered as universal primers in five Legume species: soybean, common bean, cowpea, chickpea and barrel medic. Eight of the 39 STS gave high quality sequence in all eight Legume species including P. sativum, L. japonicus and A. hypogea. Sequence comparison of the two genotypes from each species indicated that nucleotide diversity (theta) was the lowest in peanut (0.0013), cowpea (0.0011), soybean (0.0009), and Lotus (0.0003). The nucleotide diversity (theta) of pea and common bean were the highest at 0.0041 and 0.0031, respectively. The success of cross-species Legume marker development was increased if both primers were designed to coding sequence. The cross-species SNP markers discovered here can serve a useful genetic markers as well as a tool for the study of comparative Legume genomics.