Skip to main content
NCBI home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1974 Jan;1(1):171–181. doi: 10.1093/nar/1.1.171

Fingerprinting nonradioactive ribonucleic acid with the aid of polynucleotide phosphorylase

Kwok S Szeto 1, Dieter Söll 1
PMCID: PMC343333  PMID: 10793669

Abstract

We describe a method for obtaining radioactive fingerprints from nonradioactive ribonucleic acid. Fragments derived by T1 ribonuclease digestion of RNA are dephosphorylated with bacterial alkaline phosphatase. When these fragments are used as primers for the reaction of primer dependent polynucleotide phosphorylase with [α-32P]GDP in the presence of T1 ribonuclease the 3′-hydroxyl group of each fragment becomes phosphorylated. The degree of phosphorylation is reasonably uniform. The method has been applied to T1 ribonuclease digests of Escherichia coli tRNAMetf; the oligonucleotides were further analyzed by spleen phosphodiesterase digestion. In a similar manner fingerprints of pancreatic ribonuclease digests of RNA can be obtained, when [α-32P]UDP, polynucleotide phosphorylase and pancreatic ribonuclease are used.

Full text

PDF
171

Images in this article

175Image
on p.175

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Dube S. K., Marcker K. A., Clark B. F., Cory S. Nucleotide sequence of N-formyl-methionyl-transfer RNA. Nature. 1968 Apr 20;218(5138):232–233. doi: 10.1038/218232a0. [DOI] [PubMed] [Google Scholar]
  2. Dube S. K., Marcker K. A., Clark B. F., Cory S. The nucleotide sequence of N-formyl-methionyl-transfer RNA. Products of complete digestion with ribonuclease T-1 and pancreatic ribonuclease and derivation of their sequences. Eur J Biochem. 1969 Mar;8(2):244–255. doi: 10.1111/j.1432-1033.1969.tb00521.x. [DOI] [PubMed] [Google Scholar]
  3. Leder P., Singer M. F., Brimacombe R. L. Synthesis of trinucleoside diphosphates with polynucleotide phosphorylase. Biochemistry. 1965 Aug;4(8):1561–1567. doi: 10.1021/bi00884a015. [DOI] [PubMed] [Google Scholar]
  4. Mackey J. K., Gilham P. T. New approach to the synthesis of polyribonucleotides of defined sequence. Nature. 1971 Oct 22;233(5321):551–553. doi: 10.1038/233551a0. [DOI] [PubMed] [Google Scholar]
  5. SINGER M. F., HEPPEL L. A., HILMOE R. J. Oligonucleotides as primers for polynucleotide phosphorylase. J Biol Chem. 1960 Mar;235:738–750. [PubMed] [Google Scholar]
  6. Sanger F., Brownlee G. G., Barrell B. G. A two-dimensional fractionation procedure for radioactive nucleotides. J Mol Biol. 1965 Sep;13(2):373–398. doi: 10.1016/s0022-2836(65)80104-8. [DOI] [PubMed] [Google Scholar]
  7. Simsek M., Ziegenmeyer J., Heckman J., Rajbhandary U. L. Absence of the sequence G-T-psi-C-G(A)- in several eukaryotic cytoplasmic initiator transfer RNAs. Proc Natl Acad Sci U S A. 1973 Apr;70(4):1041–1045. doi: 10.1073/pnas.70.4.1041. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Székely M., Sanger F. Use of polynucleotide kinase in fingerprinting non-radioactive nucleic acids. J Mol Biol. 1969 Aug 14;43(3):607–617. doi: 10.1016/0022-2836(69)90362-3. [DOI] [PubMed] [Google Scholar]
  9. Weissmann C., Billeter M. A., Goodman H. M., Hindley J., Weber H. Structure and function of phage RNA. Annu Rev Biochem. 1973;42:303–328. doi: 10.1146/annurev.bi.42.070173.001511. [DOI] [PubMed] [Google Scholar]
  10. Yaniv M., Favre A., Barrell B. G. Structure of transfer RNA. Evidence for interaction between two non-adjacent nucleotide residues in tRNA from Escherichia coli. Nature. 1969 Sep 27;223(5213):1331–1333. doi: 10.1038/2231331a0. [DOI] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES