Polymorphism of the leghemoglobin gene in Phaseolus demonstrated by polymerase chain reaction amplification

Abstract

Genetic variance within Phaseolus vulgaris or among Phaseolus species for leghemoglobin composition may be useful in breeding for enhanced nitrogen fixation. Using primers constructed from conserved regions of the leghemoglobin gene, polymerase chain reaction was used to specifically amplify the Lba gene in total DNA samples from 17 lines and 10 species of Phaseolus. These primers are 100% homologous with the 5′- and 3′-ends of the leghemoglobin-encoding genes Lba of Phaseolus vulgaris, Lbc2 and Lbc3 of Glycine max, and 90% homologous to the G. max Lba gene. With one exception, only a single band was amplified using this approach with DNA isolated from 11 species of Phaseolus. The species of Phaseolus used in these experiments can be grouped into six classes based on the size of the amplified product which corresponded to their presumed genetic relatedness. Arranged in decreasing order by size these classes are: (1) Phaseolus lunatus and Phaseolus polystacius; (2) Phaseolus anisotrichus; (3) Phaseolus acutifolius, Phaseolus filiformis, Phaseolus angustissimus acc. # 16, and Phaseolus oligospermus; (4) Phaseolus angustissimus acc # 166, which had two major bands; (5) Phaseolus polyanthus, Phaseolus microspermus, and Phaseolus coccineus; and (6) Phaseolus vulgaris. No significant heterogeneity of amplified product within Phaseolus vulgaris was observed among 12 lines examined, but heterogeneity was observed between lines within Phaseolus acutifolius and Phaseolus angustissimus. Polymerase chain reaction amplification of conserved genes may be a useful method to facilitate the introgression of desirable genes from wild and exotic germplasm.