Biomedical subjects
Publications and source records attributed to N D Zinder.
We report a genetic analysis of a recessive chromosomal mutation of Escherichia coli K-12 that is responsibel for masking the sexual expression of an F factor that it carries. We call this new bacterial gene, which is closely linked to thr, fex, for F expression.
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The nucleotide sequence of the recognition site for the restriction-modification enzyme of Escherichia coli B (SB site) has been determined. The recognition site is a 15-nucleotide sequence consisting of the trimer 5'TGA3', followed by an 8-nucleotide domain of variable sequence, which in turn is followed by tetramer 5'TGCT3'. The sequence has no 2-fold rotational symmetry. Single base changes in the constant nucleotide domains result in the loss of sensitivity to both restriction and modification. Our data are also consistent with modification occurring by methylation of two adenine residues per SB site: one on the adenine of the trimer 5'TGA3' and the other on the complementary strand on the adenine complementary to the first thymine of the tetramer 5'TGCT3'. All nine independently isolated spontaneous mutants at the SB1 site of bacteriophage f1 are caused by a G-to-T transversion. Mutations at the SB2 site are caused by various single base changes.
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The nucleotide sequence of a region related to the initiation of the reaction in which single-stranded DNA gives rise to the replicative form (complementary strand synthesis) in bacteriophage f1 has been determined. The sequence can be drawn in an extensively base-paired structure, i.e., a single hairpin-helix 55 bases long.
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The origin and direction of synthesis in vivo of the viral and complementary strands of f1 DNA were studied by measuring the distribution of radioactivity along the genome after a short pulse of [3H]thymidine. The results indicate that the origins of replication of viral and complementary strands are located close to one another, probably both within a restriction fragment (HaeIII-G) which is about 120 bases long. Replication of both viral and complementary strands proceeds in the 5' leads to 3' overall direction. Thus, the viral strand is elongated in the counterclockwise and the complementary strand in the clockwise direction on the standard genetic map. A model is proposed in which only two mechanisms are invoked to generate all f1 DNA species: (1) the conversion of single-stranded viral DNA into double-stranded molecules and (2) the synthesis of viral single strands from double strands.
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The addition of guanidinium chloride to CsCl gradients lowers the apparent density of RNA, DNA, and hybrid polymers in such a way that all three can be banded, fractionated, and analyzed in one gradient with essentially no damage to their chemical integrity or to their biological activity.
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