PULSED-FIELD GEL-ELECTROPHORESIS ANALYSIS OF THE PHASEOLIN LOCUS REGION IN PHASEOLUS-VULGARIS

Phaseolin is the major seed storage protein of common bean (Phaseolus vulgaris is L.). It is encoded by a small multigene family of 6-9 gene s that are clustered in a single complex locus (Phs). We have construc ted a long-range restriction map of the phaseolin genomic region, incl uding the Phs locus and two flanking marker loci, D1861 and Bng060. Us ing a combination of high molecular weight DNA isolation, one- and two -dimensional pulsed-field gel electrophoresis of single and double res triction digests followed by Southern hybridization, and PCR analysis of individual fragments, we found that: (i) the maximum size of the Ph s locus is 190 kb, (ii) the Phs locus may have increased in size durin g the evolution of P. vulguris, (iii) the genomic region marked by D18 61-Phs-Bng060 spans 5 cM, which corresponds to a maximum of 1.9 Mb, an d (iv) the Phs locus could be oriented with respect to the two adjacen t markers. Further progress in determining the gene arrangement in the Pits locus will require cloning and analysis of large DNA fragments c ontaining phaseolin genes via BAC libraries.