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目的克隆汉族人甘露聚糖结合凝集素(MBL)全长基因。方法提取汉族人白细胞基因组DNA,高保真PCR扩增MBL基因,应用TA克隆方法将PCR产物与载体pCR-XL-TOPO连接,经DNA测序后利用软件DNAStar进行分析。结果MBL基因反向插入pCR-XL-TOPO,全长6321bp,与GenBank收录的基因序列相比,有17个碱基不同,其中仅1个位于编码区(外显子4),但编码的氨基酸没有改变,且该碱基恰与GenBank收录的人MBL cDNA一致,此序列已被GenBank收录,登录号EU596574。结论成功克隆了汉族人MBL基因全长序列,其结构基因的基因型属于野生型,这为研究MBL基因非编码区对MBL表达的影响奠定了基础。
Objective To clone full length Han Chinese mannan binding lectin (MBL) gene. Methods Genomic DNA of Han white blood cells was extracted. The MBL gene was amplified by high-fidelity PCR. The PCR products were ligated with pCR-XL-TOPO by TA cloning method and analyzed by DNAStar software after DNA sequencing. RESULTS: The MBL gene was inserted into pCR-XL-TOPO in the reverse direction with a total length of 6321bp. Compared with the GenBank-based gene sequence, MBL contained 17 different bases, of which only 1 locates in the coding region (exon 4) The amino acid has not changed, and the base exactly matches with the human MBL cDNA collected from GenBank. This sequence has been deposited with GenBank and has accession number EU596574. Conclusion The full-length MBL gene sequence was successfully cloned from Han people and the genotypes of the structural genes were wild type, which laid the foundation for studying the effect of non-coding region of MBL on the expression of MBL.