Expression of human and Chinese hamster hypoxanthine-guanine phosphoribosyltransferase cDNA recombinants in cultured Lesch-Nyhan and Chinese hamster fibroblasts.
Expression of human and Chinese hamster hypoxanthine-guanine phosphoribosyltransferase cDNA recombinants in cultured Lesch-Nyhan and Chinese hamster fibroblasts.
复制标题
人和中国仓鼠次黄嘌呤鸟嘌呤磷酸核糖基转移酶 cDNA 重组体在培养的 Lesch-Nyhan 和中国仓鼠成纤维细胞中的表达。
DOI：
10.1016/s0021-9258(17)44533-9
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发表时间：
1983
期刊：
影响因子：
--
通讯作者：
C. Caskey
中科院分区：
文献类型：
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作者：
John BrennandS;David S. Konecki;C. Caskey
结果表明，次黄嘌呤鸟嘌呤磷酸核糖基转移酶(HPRT)基因在中国仓鼠和人成纤维细胞中均可表达，内源性基因产物缺陷，转化子的正常生长水平。表达所需的所有元件都存在于pBR322衍生的含有HPRT cDNA编码序列和逆转录病毒长末端重复序列的质粒中。这些分子在所研究的两个物种中都起作用，至少在中国仓鼠的转化体中，在单拷贝水平上是有效的。尽管质粒中存在内含子序列和多聚腺苷酸化信号的影响尚未得到评估，但这些研究表明，在培养的哺乳动物细胞中，两者都不是表达HPRT基因序列的绝对要求。我们描述了构建含有野生型人或中国仓鼠HPRT cDNA序列的重组载体，并将其与逆转录病毒LTR连接，在HPRT缺失的V79和Lesch-Nyhan成纤维细胞中赋予HPRT+表型。高频率分离到稳定和不稳定的HPRT基因和蛋白的转化子。
The results presented in this communication demonstrate that hypoxanthine-guanine phosphoribosyltransferase (HPRT) cDNA can be expressed in both Chinese hamster and human fibroblasts deficient in the endogenous gene product at levels permitting normal growth of the transformants. All the elements necessary for this expression are present in a pBR322-derived plasmid containing HPRT cDNA coding sequence and a retroviral long terminal repeat. These molecules function in both species investigated and, at least in the case of the Chinese hamster transformants, are efficient at the single copy level. Although the effects of the presence of intron sequences and a polyadenylation signal within the plasmids have yet to be evaluated, these studies demonstrate that neither is an absolute requirement for expression of HPRT cDNA sequences in cultured mammalian cells. We describe the construction of recombinant plasmids containing wild type human or Chinese hamster HPRT cDNA sequences in tandem with a retroviral LTR which confer the HPRT+ phenotype in HPRT-deficient V79 and Lesch-Nyhan fibroblasts. Both stable and unstable transformants, that expressed HPRT mRNA and protein, were isolated at high frequency.