Duvoisin R M, Hayzer D J, Belin D, Jaton J C
Department of Medical Biochemistry, Faculty of Medicine, University Medical Center, Geneva, Switzerland.
J Immunol. 1988 Sep 1;141(5):1596-601.
Southern blot analyses of germ-line DNA obtained from rabbits expressing lambda chains of C7 and/or C21 allotypes were performed with a rabbit C lambda region-specific probe; a 12-kbp EcoRI- and a 2-kbp BamHI-hybridizing fragment were detected only in the DNA from rabbits expressing the C21 allotype. The 12-kbp EcoRI fragment was cloned and shown to contain two C lambda region-encoding genes in the same orientation. Each is preceded by a J lambda gene segment. Nonamer-12-bp spacer-heptamer recombination signal sequences were found 5' of each J lambda segment, and splicing signals were identified at the 3' ends of the J lambda segments and the 5' ends of the corresponding C lambda genes. The C lambda 5 gene, which exhibits a sequence identical with that found in several cDNA clones, is carried by the 2-kbp BamHI fragment missing from the genomic DNA of rabbits which do not express the C21 allotype. The second C lambda gene, C lambda 6, lies 3' of C lambda 5, in a 1.6-kbp BamHI fragment which is present in genomic DNAs of all tested rabbits, irrespective of their phenotype. Its sequence is identical with that found in one cDNA clone and differs from that of C lambda 5 in 17 base positions resulting in four amino acid substitutions. A fragment of a cDNA, with a J-C region sequence identical with that encoded by the J lambda 5-C lambda 5 gene pair, was subcloned into a plasmid expression vector. The resulting polypeptide product could be specifically immunoprecipitated by anti-C21 but not anti-C7 alloantisera, showing that some, if not all, C21 allotopes are encoded by the C lambda 5 gene. In contrast, the C lambda 6 gene product was not precipitable, either by anti-C7 or by anti-C21 alloantisera, although it was readily immunoprecipitated by a goat anti-rabbit lambda chain antiserum.
用兔Cλ区域特异性探针,对表达C7和/或C21同种异型λ链的兔的种系DNA进行Southern印迹分析;仅在表达C21同种异型的兔的DNA中检测到一个12kbp的EcoRI杂交片段和一个2kbp的BamHI杂交片段。克隆了12kbp的EcoRI片段,发现其包含两个同向的Cλ区域编码基因。每个基因之前都有一个Jλ基因片段。在每个Jλ片段的5'端发现了九聚体-12bp间隔区-七聚体重组信号序列,并且在Jλ片段的3'端和相应Cλ基因的5'端鉴定出剪接信号。Cλ5基因,其序列与在几个cDNA克隆中发现的序列相同,由不表达C21同种异型的兔的基因组DNA中缺失的2kbp BamHI片段携带。第二个Cλ基因,Cλ6,位于Cλ5的3'端,在一个1.6kbp的BamHI片段中,该片段存在于所有测试兔的基因组DNA中,无论其表型如何。其序列与在一个cDNA克隆中发现的序列相同,与Cλ5的序列在17个碱基位置不同,导致四个氨基酸取代。一个cDNA片段,其J-C区域序列与由Jλ5-Cλ5基因对编码的序列相同,被亚克隆到一个质粒表达载体中。所得的多肽产物可以被抗C21但不能被抗C7同种异型抗血清特异性免疫沉淀,表明一些(如果不是全部)C21同种异型抗原决定簇由Cλ5基因编码。相反,Cλ6基因产物既不能被抗C7也不能被抗C21同种异型抗血清沉淀,尽管它很容易被山羊抗兔λ链抗血清免疫沉淀。