Molecular approaches to eucaryotic genetic systems / edited by Gary Wilcox, John Abelson, C. Fred Fox.

Date:
1977
Catalogue details

Licence: Attribution-NonCommercial 4.0 International (CC BY-NC 4.0)

Credit: Molecular approaches to eucaryotic genetic systems / edited by Gary Wilcox, John Abelson, C. Fred Fox. Source: Wellcome Collection.

    19/472 (page 1)
    NUCLEOTIDE SEQUENCE OF BACTERIOPHAGE 0X174 DNA John C. Fiddes MRC Laboratory of Molecular Biology Hills Road, Cambridge CB2 2QH, England ABSTRACT. The plus and minus technique has been used by Sanger et al. (1) to obtain an almost complete nucleotide sequence for the DNA of the single stranded bacteriophage 0X174. The locations of the initiation and termination sites for the nine known genes and the extent of the intercistronic regions were determined. Two pairs of genes have been demon¬ strated to overlap, with the nucleotide sequence being read in different phases. Gene В is thus entirely contained within gene A, and likewise gene E is within gene D. In this article some features of the 0X174 nucleotide sequence are discussed. INTRODUCTION The plus and minus technique (2) for DNA nucleotide sequence analysis is based on primed synthesis of DNA by DNA polymerase I. This is usually accomplished with single stranded DNA as template annealed to a restriction enzjmie fragment as specific primer. The details of the method will not be discussed here. The technique is ideally suited for studying single stranded DNA bacteriophages such as 0X174 since the viral DNA provides a naturally occurring single stranded DNA template. The 0X174 complementary DNA strand can also be used since it is easily separable from the viral DNA strand when 0X174 double stranded DNA is centrifuged in alkaline caesium chloride gradients. Detailed cleavage maps of the 0X174 replicative form (RF) DNA are known for a range of restric¬ tion enzymes (3-9) and these have been correlated with the 0X174 genetic map by the marker rescue method (10-12). Another reason for sequencing 0X174 DNA is that the biology of the phage has been studied extensively (see review by Denhardt, ref. 13). 0X174 has one of the smallest known DNA genomes containing only about 5400 nucleotides. Nine genes have been identified by recombination and complementa¬ tion analysis and protein products have been related to them. The combined molecular weights of these proteins more than account for the coding capacity of the phage genome. The locations of the three promoters and possible sites for the termination of transcription (14-16) and the position of the 1