人牙本质基质蛋白1cDNA序列的克隆

Cloning and partially sequencing of human dentin matrix protein 1 encoding mature protein

目的：克隆人牙本质基质蛋白１（ＤＭＰ１）成熟肽编码区基因片段。方法：用异硫氰酸胍一步法从引产胎儿牙乳头组织中抽提总ＲＮＡ，用Ｏｌｉｇｏ（ｄｔ）作引物逆转录合成牙乳头ｃＤＮＡ，然后利用ＰＣＲ方法，从ｃＤＮＡ中扩增出人ＤＭＰ１成熟区的基因片段（约１．８ｋｂｐ），将所得基因片段插入ｐＢｌｕｅｓｃｒｉｐｔ质粒载体，转化到大肠杆菌ＸＬ１－Ｂｌｕｅ后挑选阳性克隆，提取重组质粒ＤＮＡ，通过酶切分析和核苷酸序列分析鉴定阳性克隆。结果：酶切图谱和部分序列分析结果与国外文献报道一致。结论：克隆到人ＤＭＰ１成熟肽编码区基因片段。

Aim: Cloning and partially sequencing of human dentin matrix protein 1 (DMP1) encoding mature protein. Methods: In the study, total RNA was extracted from the tooth germs of a legally aborted embryo by acid guanidinium thiocyanata-phenol-chloroform method, the desired DNA product was obtained from the total RNA by RT-PCR with the primers including Oligo(dt) and two gene specific primers. The segment (about 1.8 kbp) was inserted into pBluescript vector and the interesting plasmid was transformed into E. Coli host strain XL1-Blue. The double-stranded DNA of the positive clone was analyzed by restriction endonuclease mapping and DNA sequencing. Results: The restriction endonuclease map and sequence of human DMP1 encoding mature protein were consistent with those of the references published. Conclusion: The human DMP1 cDNA encoding mature protein was obtained for further research.