Sequence analysis of low-molecular-weight RNAs by the use of non-radioactive labeling.
A new procedure for non-radioactive labeling of the 3'-terminal -OH was developed to facilitate the sequencing works on low-molecular-weight RNAs. At first, biotinylated pCp (cytidine 3',5'-bisphosphate) was synthesized and ligated to the 3'-terminal of yeast tRNA(Phe) with the aid of RNA ligase. Although the labeling efficiency was high enough, this labeling resulted in a lack of the sequence ladders around the 3'-terminal 10 nucleotides. A longer oligonucleotide, p(dC)(dT)16-Biotin was designed to overcome this defect and proved to be a better non-radioactive 3'-labeling probe.
Matsuki
T
T
Department of Biological Sciences, Faculty of Bioscience and Biotechnology, Tokyo Institute of Technology, Yokohama, Japan.
Ito
R
R
Yokogawa
T
T
Nishikawa
K
K
D001710:Biotin; D008163:Luminescent Measurements; D008970:Molecular Weight; D011130:RNA Ligase (ATP) / Q000737:chemistry; D012360:RNA, Transfer, Phe / Q000737:chemistry*; D012441:Saccharomyces cerevisiae / Q000235:genetics; D017423:Sequence Analysis, RNA / Q000379:methods*
D012360:RNA, Transfer, Phe; D001710:Biotin; D011130:RNA Ligase (ATP)