摘要
目的：克隆小鼠牙本质涎蛋白（ＤＳＰ）成熟肽编码区基因。方法：用异硫氰酸胍一步法从昆明新生小鼠磨牙牙胚组织中抽提总ＲＮＡ，用Ｏｌｉｇｏ（ｄｔ）作引物逆转录合成牙胚ｃＤＮＡ，然后利用ＰＣＲ方法，从ｃＤＮＡ中扩增出小鼠ＤＳＰ成熟区的基因片段（约１．４ｋｂｐ），将所得基因片段插入ｐＢＳ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，通过限制性酶切和核苷酸序列分析鉴定阳性克隆。结果：重组质粒ｐＢＳ－ＤＳＰ的酶切图谱和部分序列分析结果与国外文献报道一致。结论：克隆到小鼠ＤＳＰ成熟肽编码区基因，正在进行该基因的表达和活性鉴定。
Aim: Cloning and partially sequencing of mouse dentin sialoprotein (DSP) encoding mature protein. Methods: In the study, total RNA was extracted from the tooth germs of newborn mouse 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 1.4 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 mouse DSP encoding mature protein were consistent with those of the references appeared. Conclusion: the mouse DSP cDNA encoding mature protein was obtained for further study.
出处
《牙体牙髓牙周病学杂志》
CAS
1999年第1期39-41,共3页
Chinese Journal of Conservative Dentistry
基金
国家自然科学基金
上海市博士后科研资助
中科院上海细胞所开放实验室资助
