生物
T7 RNA聚合酶
寡核苷酸
终端(太阳能)
核糖核酸
聚合酶
RNA聚合酶Ⅲ
终止因子
抄写(语言学)
克莱诺碎片
RNA依赖性RNA聚合酶
反终止
小核RNA
RNA聚合酶
分子生物学
DNA
遗传学
计算生物学
基因
噬菌体
核酸外切酶
电离层
物理
哲学
语言学
大肠杆菌
天文
出处
期刊:Gene
[Elsevier]
日期:1988-12-01
卷期号:72 (1-2): 75-89
被引量:85
标识
DOI:10.1016/0378-1119(88)90129-1
摘要
This communication presents an overview of the methods for the synthesis of RNA with virtually any desired sequence. The use of transcription vectors is a powerful and convenient approach, if the cloned gene of interest has restriction sites at the proper positions. To overcome these limitations, two methods were developed where chemically synthesized oligodeoxynucleotides (oligos) were applied to define the 3' and 5' termini of the chosen transcripts. Both approaches use cloned genes and the template DNA is synthesized with DNA polymerase I (Klenow fragment). Consequently, there are no size limitations for the synthesized RNAs. For short transcripts, the entire template DNA (including the promoter sequence) can be synthesized chemically and any desired RNA sequence is possible. Recently, it was shown that even oligos without any promoter sequence can be used as template DNA for RNA polymerases. Experimental data are presented for two approaches. The first example is the synthesis of template DNA for T7 RNA polymerase where two oligos (initiator and terminator) define the beginning and end of transcripts from a cloned gene. The second example is the use of simple oligos as templates for RNA polymerases. The major problem encountered was the inaccurate transcription termination, which resulted in one or two additional nucleotides beyond the encoded sequence.
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