[Economical sequencing of DNA with terminators].
We describe several improvements of chain-termination DNA sequencing procedure of Sanger et al. For template preparation we use 0.3 ml cultures of M13 clones, grown in standard 1,5 ml polypropylene tubes. The sequencing experiment differs from the previously described by the use of deoxyNTP, labelled with phosphorus-33 (a low energy isotope with a half-life of 25 days, commercially produced in the USSR), and by a "quasi-end labelling" reaction, preceding the DNA synthesis in the presence of dideoxyNTPs. The combination of the phosphorus-33 and the quasi-end labelling produces very sharp sequencing ladders, that equal or exceed in quality those obtained with sulphur-35, and only an overnight exposure with a conventional X-ray film is required. The use of plastic tubes for bacterial growth and the 60-well microchambers for carrying out sequencing reactions results in substantial saving of time and cost in routine "middle scale" sequencing (both types of plasticware are produced in the USSR).
Kraev
A S
AS
Mironov
V N
VN
D003001:Cloning, Molecular; D003365:Costs and Cost Analysis; D004247:DNA / Q000737:chemistry*; D009710:Nucleotide Mapping / Q000379:methods*; D016133:Polymerase Chain Reaction; D013728:Terminator Regions, Genetic
D004247:DNA
Ekonomichnoe sekvenirovanie DNK s terminatorami.