Coleman Tricia M, Wang Guocan, Huang Faqing
Department of Chemistry and Biochemistry, University of Southern Mississippi, Hattiesburg, MS 39406-5043, USA.
Nucleic Acids Res. 2004 Jan 15;32(1):e14. doi: 10.1093/nar/gnh007.
Transcription from the commonly used GTP- initiating T7 class III promoter phi6.5 frequently produces heterogeneous RNA at both 3' and 5' ends. We demonstrate here that RNA transcripts from the T7 class II promoter phi2.5 have superior 5' homogeneity over those from the phi6.5 promoter, with comparable total RNA yields. The overall homogeneity of RNA transcripts is improved to different degrees depending on RNA sequences, although transcription under phi2.5 does not affect the 3' heterogeneity of RNA. In combination with 3' RNA trimming by DNAzymes or ribozymes, this ATP- initiated transcription system based on the T7 phi2.5 promoter can provide excellent quality of RNA for applications requiring a high degree of RNA size homogeneity.
从常用的启动GTP的T7 III类启动子phi6.5转录时,3'和5'末端常常产生异质性RNA。我们在此证明,来自T7 II类启动子phi2.5的RNA转录本在5'端的同质性优于来自phi6.5启动子的转录本,且总RNA产量相当。尽管在phi2.5启动子下转录不会影响RNA的3'端异质性,但RNA转录本的整体同质性会根据RNA序列不同程度地得到改善。结合通过脱氧核酶或核酶进行的3'端RNA修剪,这种基于T7 phi2.5启动子的ATP启动转录系统可为需要高度RNA大小同质性的应用提供优质的RNA。