A HUMAN BETA-ACTIN EXPRESSION VECTOR SYSTEM DIRECTS HIGH-LEVEL ACCUMULATION OF ANTISENSE TRANSCRIPTS
A HUMAN BETA-ACTIN EXPRESSION VECTOR SYSTEM DIRECTS HIGH-LEVEL ACCUMULATION OF ANTISENSE TRANSCRIPTS
复制标题
DOI：
10.1073/pnas.84.14.4831
复制
发表时间：
1987-07-01
影响因子：
11.1
通讯作者：
KEDES, L
中科院分区：
文献类型：
--
作者：
GUNNING, P;LEAVITT, J;KEDES, L
我们已经构建了由人β-半乳糖苷酶的3种酶组成的哺乳动物表达载体。肌动蛋白基因5 ″侧翼序列加上5 ″非翻译区和间插序列1，其在3 ″剪接位点处连接到短DNA多接头序列，所述短DNA多接头序列含有独特的Sal I、HindIII和BamHI限制性内切酶位点，随后是猿猴病毒40（SV 40）多腺苷酸化信号。还产生了含有从pSVgpt或pSV 2neo获得的选择标记的两种衍生物。我们发现，该载体的启动子活性与多种人类和啮齿动物细胞中的SV 40早期启动子活性一样大或更大。该载体用于产生γ-肌动蛋白和β-人成纤维细胞系中的微管蛋白反义转录本。反义转录物积累到与高度丰富的γ转录物相当的水平。肌动蛋白和β-微管蛋白mRNA。
We have constructed a mammalian expression vector consisting of 3 kilobases of the human .beta.-actin gene 5'' flanking sequence plus 5'' untranslated region and intervening sequence 1 linked at the 3'' splice site to a short DNA polylinker sequence containing unique Sal I, HindIII, and BamHI restriction endonuclease sites followed by a simian virus 40 (SV40) polyadenylylation signal. Two derivatives containing the selection markers obtained from pSVgpt or pSV2neo, were also generated. We find that the promoter activity of this vector is as great or greater than that of the SV40 early promoter in a variety of human and rodent cells. The vector was used to generate .gamma.-actin and .beta.-tubulin antisense transcripts in human fibroblast cell lines. The antisense transcripts accumulate to levels comparable with that of the highly abundant .gamma.-actin and .beta.-tubulin mRNAs.