Jaspers M, Marynen P, Aly M S, Cuppens H, Hilliker C, Cassiman J J
Center for Human Genetics, University of Leuven, Belgium.
Somat Cell Mol Genet. 1991 Sep;17(5):505-11. doi: 10.1007/BF01233174.
The alpha 2 subunit of the VLA-2 receptor (CD49B) was mapped to human chromosome 5 by several independent approaches. First, the expression of the alpha 2 subunit at the protein level was investigated in a panel of human-mouse hybrid cell lines. Cell surface expression was detected by indirect immunofluorescence with monoclonal anti-alpha 2 antibody 12F1. Intracellular alpha 2 antigen was detected by immunostaining of whole cell extracts or of immunoprecipitated 12F1 antigen with the monoclonal antibodies 3H8 and 5C5. Second, the presence of human genomic alpha 2 sequences in the panel of human-mouse hybrids was detected by PCR, using primers derived from the published alpha 2 cDNA sequence. The specificity of the amplification product was shown by direct sequencing. The results of the PCR study were confirmed by amplifying a CD14 gene fragment, known to map to chromosome 5. Finally, in situ hybridization with a 3H-labeled 1040-bp cDNA probe, also obtained by PCR, confirmed and refined the localization of CD49B on chromosome 5 at q23-31.
通过几种独立的方法,将VLA - 2受体(CD49B)的α2亚基定位到了人类5号染色体上。首先,在一组人 - 鼠杂交细胞系中研究了α2亚基在蛋白质水平的表达。通过用单克隆抗α2抗体12F1进行间接免疫荧光检测细胞表面表达。通过用单克隆抗体3H8和5C5对全细胞提取物或免疫沉淀的12F1抗原进行免疫染色来检测细胞内α2抗原。其次,使用从已发表的α2 cDNA序列衍生的引物,通过PCR检测人 - 鼠杂交细胞系中人类基因组α2序列的存在。通过直接测序显示扩增产物的特异性。通过扩增已知定位于5号染色体的CD14基因片段,证实了PCR研究的结果。最后,用同样通过PCR获得的3H标记的1040bp cDNA探针进行原位杂交,证实并细化了CD49B在5号染色体q23 - 31处的定位。