Tabor S
Harvard Medical School, Boston, Massachusetts, USA.
Curr Protoc Mol Biol. 2001 May;Chapter 3:Unit3.7. doi: 10.1002/0471142727.mb0307s00.
Reverse transcriptases are derived from retroviruses which use them to make DNA copies of their RNA genomes. They are multifunctional enzymes, but they are mainly used as RNA-directed DNA polymerases. Specifically, deoxyoligonucleotides (either oligo(dT) polymers, a collection of random DNA sequences, or a specific sequence) are used as primers for extension on RNA (usually messenger RNA) templates. The DNA synthesized from the RNA template is referred to as complementary DNA (cDNA). This unit describes appropriate reaction conditions along with applications such as synthesizing cDNA for insertion into bacterial cloning vectors, filling in and labeling the 3'termini of DNA fragments with 5' protruding ends, and its use for DNA sequencing in place of Klenow fragment.
逆转录酶来源于逆转录病毒,逆转录病毒利用它们将其RNA基因组复制成DNA。它们是多功能酶,但主要用作RNA指导的DNA聚合酶。具体而言,脱氧寡核苷酸(要么是寡聚(dT)聚合物、一组随机DNA序列,要么是特定序列)用作在RNA(通常是信使RNA)模板上进行延伸的引物。从RNA模板合成的DNA称为互补DNA(cDNA)。本单元描述了合适的反应条件以及一些应用,如合成用于插入细菌克隆载体的cDNA、填补和标记具有5'突出末端的DNA片段的3'末端,以及用其替代Klenow片段用于DNA测序。