Identification of molecular markers in soybean comparing RFLP, RAPD and AFLP DNA mapping techniques
1996
Lin, J.J. | Kuo, J. | Ma, J. | Saunders, J.A. | Beard, H.S. | MacDonald, M.H. | Kenworthy, W. | Ude, G.N. | Matthews, B.F.
Restriction fragment length polymorphism (RFLP), random amplified polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP) were used on identical soyabean germplasm to compare their ability to identify markers in the development of a genetic linkage map. Polymorphisms present in 14 soyabean cultivars were demonstrated using all 3 techniques. AFLP, a PCR-based technique, was able to identify multiple polymorphic bands in a denaturing gel using 60 of 64 primer pairs tested. AFLP relies on primers designed in part on sequences for endonuclease restriction sites and on three selective nucleotides. The 60 diagnostic primer pairs tested for AFLP analysis each distinguished on average 6 polymorphic bands. Using specific primers designed for soyabean from EcoRI and MseI restriction site sequences and three selective nucleotides, as many as 12 polymorphic bands per primer could be obtained with AFLP techniques. Only 35% of the RAPD reactions identified a polymorphic band using the same soyabean cultivars, and in those positive reactions, typically only one or two polymorphic bands per gel were found. Identification of polymorphic bands using RFLP techniques was the most cumbersome, because Southern blotting and probe hybridization were required. Over 50% of the soyabean RFLP probes examined failed to distinguish even a single polymorphic band, and the RFLP probes that did distinguish polymorphic bands seldom identified more than one polymorphic band. It is concluded that, of the 3 techniques tested, AFLP is the most useful.
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