Utilization of a Potentially Universal Downstream Primer in the Rapid Identification and Characterization of Vβ Genes from Two New Human Vλ Gene Families

Polymerase chain reaction (PCR) has increased dramatically the speed of cloning and characterizing numerous genes. However, its application to identifying and analysing new germline Ig-variable (V) gene families has been hampered by the lack of sequence information in the downstream flanking regions of the concerned V genes, which are deleted during V(D)J rearrangements. To circumvent this problem, the possibility was explored that a degenerate downstream primer may be used in conjunction with a specific upstream primer, to clone members of new V lambda gene families, as much less is known about V lambda genes than Vh and Vk genes in humans. Firstly the feasibility and the specificity of a degenerate primer was examined by comparing it with an established downstream primer in amplifying known V lambda 1 genes. The results were positive. Thus, the degenerate primer was used to clone and characterize germline V lambda genes of the recently defined V lambda 8 and V lambda 9 gene families. This current strategy may help speed up the identification and characterization of all human V lambda genes. Moreover, a similar strategy can be applied to identify and characterize rapidly new V genes of either known or unknown Ig and T-cell receptor V gene families.