Molecular cloning of Pst I fragments from rat double stranded thyroglobulin complementary DNA.
Molecular cloning of Pst I fragments from rat double stranded thyroglobulin complementary DNA.
复制标题
大鼠双链甲状腺球蛋白互补 DNA 的 Pst I 片段的分子克隆。
DOI：
10.1016/0006-291x(80)91381-9
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发表时间：
1980
影响因子：
3.1
通讯作者：
Vassart,G
中科院分区：
文献类型：
--
作者：
Brocas,H;Christophe,D;VanHeuverswijn,B;Scherberg,N;Vassart,G
从丙基硫氧嘧啶长期治疗的大鼠甲状腺制备甲状腺球蛋白mRNA。使用禽成髓细胞瘤病毒逆转录酶合成双链互补DNA，并用内切核酸酶Pst I进行限制性内切。将限制性片段连接到质粒pBR 322的独特Pst I位点，并将所得重组DNA用于转化大肠杆菌以获得四环素抗性。携带重组质粒的菌落（1852个中的42个）可分为含有320、550和640个碱基对插入物的三类。从每个大小类别中选择一个克隆，并在正翻译测定中证明其质粒DNA结合功能活性大鼠甲状腺球蛋白mRNA的能力。这三个片段共占甲状腺球蛋白结构基因序列的20%左右。
Thyroglobulin mRNA was prepared from thyroid glands of rats chronically treated with propylthiouracil. The double stranded complementary DNA was synthesized using Avian Myeloblastosis Virus reverse transcriptase and subjected to restriction with the endonuclease Pst I. The restriction fragments were ligated into the unique Pst I site of the plasmid pBR 322 and the resulting recombinant DNA was used to transform E.coli to tetracyclin resistance. The colonies harboring recombinant plasmids (42 out of 1852) could be classified into three categories containing 320, 550 and 640 base pair inserts. One clone from each size class was selected and the ability of their plasmid DNA to bind functionaly active rat thyroglobulin mRNA was demonstrated in a positive translation assay. Altogether the three cloned DNA fragments represent about 20% of thyroglobulin structural gene sequence.