Van Coillie E, Fiten P, Nomiyama H, Sakaki Y, Miura R, Yoshie O, Van Damme J, Opdenakker G
Rega Institute for Medical Research, Laboratory of Molecular Immunology, University of Leuven, Belgium.
Genomics. 1997 Mar 1;40(2):323-31. doi: 10.1006/geno.1996.4594.
Monocyte chemotactic proteins (MCPs) form a subfamily of chemokines that recruit leukocytes to sites of inflammation and that may contribute to tumor-associated leukocyte infiltration and to the antiviral state against HIV infection. With the use of degenerate primers that were based on CC chemokine consensus sequences, the known MIP-1 alpha/LD78 alpha, MCP-1, and MCP-3 genes and the previously unidentified eotaxin and MCP-2 genes were isolated from a YAC contig from human chromosome 17q11.2. The amplified genomic MCP-2 fragment was used to isolate an MCP-2 cosmid from which the gene sequence was determined. The MCP-2 gene shares with the MCP-1 and MCP-3 genes a conserved intron-exon structure and a coding nucleotide sequence homology of 77%. By Northern blot analysis the 1.0-kb MCP-2 mRNA was predominantly detectable in the small intestine, peripheral blood, heart, placenta, lung, skeletal muscle, ovary, colon, spinal cord, pancreas, and thymus. Transcripts of 1.5 and 2.4 kb were found in the testis, the small intestine, and the colon. The isolation of the MCP-2 gene from the chemokine contig localized it on YAC clones of chromosome 17q11.2, which also contain the-eotaxin, MCP-1, MCP-3, and NCC-1/MCP-4 genes. The combination of using degenerate primer PCR and YACs illustrates that novel genes can efficiently be isolated from gene cluster contigs with less redundancy and effort than the isolation of novel ESTs.
单核细胞趋化蛋白(MCPs)构成趋化因子的一个亚家族,可将白细胞募集到炎症部位,并可能促进肿瘤相关白细胞浸润以及对抗HIV感染的抗病毒状态。利用基于CC趋化因子共有序列的简并引物,从人染色体17q11.2的一个酵母人工染色体(YAC)重叠群中分离出已知的MIP-1α/LD78α、MCP-1和MCP-3基因以及先前未鉴定的嗜酸性粒细胞趋化因子和MCP-2基因。扩增的基因组MCP-2片段用于分离一个MCP-2黏粒,并测定其基因序列。MCP-2基因与MCP-1和MCP-3基因具有保守的内含子-外显子结构,编码核苷酸序列同源性为77%。通过Northern印迹分析,在小肠、外周血、心脏、胎盘、肺、骨骼肌、卵巢、结肠、脊髓、胰腺和胸腺中主要可检测到1.0 kb的MCP-2 mRNA。在睾丸、小肠和结肠中发现了1.5 kb和2.4 kb的转录本。从趋化因子重叠群中分离出MCP-2基因,将其定位在染色体17q11.2的YAC克隆上,该克隆还包含嗜酸性粒细胞趋化因子、MCP-1、MCP-3和NCC-1/MCP-4基因。使用简并引物PCR和YACs相结合的方法表明,与分离新的ESTs相比,从基因簇重叠群中可以更高效地分离出新基因,且冗余度更低、工作量更小。