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Cleavage maps of the filamentous bacteriophages M13, fd, fl, and ZJ/2
Abstract:
The replicative form DNAs of bacteriophage M13, fd, f1, and ZJ/2 were found to be sensitive to cleavage by the restriction endonucleases endoR-HapII, endoR-HaeII, endoR-HaeIII, endoR-HindII, endoR-AluI, endoR-Hha, and endoR-Hinf. With respect to M13 DNA the number of cleavage sites varied from 21 for endoR-Hinf, 18 for endoR-AluI, 15 for endoR-Hha, 13 for endoR-HapII, 10 for endoR-HaeIII, 3 for endoR-HaeII, to only a single site for endoR-HindII. In contrast to M13, fd and f1, the ZJ/2 DNA molecule was not cleaved by the endoR-HindII endonuclease. No cleavage site on either phage DNA was detected for the endonucleases endoR-Hsu, endoR-EcoRI and endoR-Sma. When compared with M13 DNA, several differences were noted in the number and size of cleavage products obtained with DNA of phage fd, f1, and ZJ/2. From the results of these analyses, using the M13 enzyme cleavage maps as a reference, the endoR-HapII, endoR-HaeII, endoR-HaeIII, endoR-HindII and endoR-AluI maps of phage fd, f1, and ZJ/2 could be constructed. As is expected for very closely related phages, the enzyme cleavage patterns exhibit a high degree of homology. Phage f1 and ZJ/2 are most related since an identical pattern was obtained with seven different restriction endonucleases. Evidence is provided also that f1 is more similar to M13 than to fd. Furthermore, characteristic differences exist within the endoR-Hinf enzyme cleavage pattern of all the four phages tested. Digestion of phage DNA with this enzyme, therefore, provides a new and sensitive method of distinguishing these closely related filamentous coliphages .
Insights
Restriction endonucleases cleave filamentous coliphage DNA, revealing genetic similarities and differences. Digestion with endoR-Hinf offers a sensitive method to distinguish closely related phages like M13, fd, f1, and ZJ/2.
Area of Science:
- Molecular Biology
- Virology
- Genetics
Background:
- Bacteriophages M13, fd, f1, and ZJ/2 are closely related filamentous phages.
- Understanding their genetic relatedness is crucial for molecular biology applications.
Purpose of the Study:
- To analyze the susceptibility of these phage DNAs to various restriction endonucleases.
- To construct restriction maps for comparative analysis.
- To identify sensitive methods for differentiating these phages.
Main Methods:
- Digestion of replicative form DNA from phages M13, fd, f1, and ZJ/2 with a panel of restriction endonucleases.
- Analysis of cleavage products to determine the number and location of restriction sites.
- Construction of restriction maps based on M13 DNA as a reference.
Main Results:
- All tested phage DNAs were sensitive to endoR-HapII, endoR-HaeII, endoR-HaeIII, endoR-HindII, endoR-AluI, endoR-Hha, and endoR-Hinf, with varying numbers of cleavage sites.
- Phage ZJ/2 DNA was not cleaved by endoR-HindII, unlike M13, fd, and f1.
- Restriction maps were constructed for fd, f1, and ZJ/2 DNA using M13 maps as a reference.
- Phages f1 and ZJ/2 showed the highest relatedness with identical patterns for seven endonucleases.
- endoR-Hinf digestion provided characteristic differences, enabling sensitive distinction among the four phages.
Conclusions:
- The restriction endonuclease cleavage patterns reveal a high degree of homology among these filamentous coliphages.
- Phage f1 is more similar to M13 than to fd.
- endoR-Hinf digestion is a valuable tool for distinguishing between these closely related phages.