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Physical map of pColA-CA31, an amplifiable plasmid, and location of colicin A structural gene
Abstract:
Evidence showing that the plasmic ColA, derived from strain CA31[pColA] can be amplified in the presence of chloramphenicol is presented. This plasmid has been purified and its Mr-value has been found to be 4.6 X 10(6) or 7 kb. Twelve cleavage sites have been mapped in pColA by using single and double restriction endonuclease digestions. These sites were ordered in relation to the single HindIII site. The other restriction endonucleases used were, respectively, SmaI, AvaI, PstI and HincII. Establishment of the map was helped by hybridization of pColA endonuclease digest products with 32P-labeled colicin A-mRNA. The structural gene for colicin A was contained in a 2.17-kb HincII fragment.
Insights
The ColA plasmid (pColA) can be amplified with chloramphenicol. Researchers mapped its restriction sites and identified the colicin A structural gene within a specific DNA fragment.
Area of Science:
- Molecular Biology
- Genetics
- Biochemistry
Background:
- The ColA plasmid (pColA) is a genetic element found in certain bacterial strains.
- Understanding plasmid structure and gene organization is crucial for molecular biology research.
Purpose of the Study:
- To characterize the physical properties of the ColA plasmid.
- To create a restriction endonuclease map of pColA.
- To locate the structural gene for colicin A on the plasmid.
Main Methods:
- Plasmid amplification in the presence of chloramphenicol.
- Purification and molecular weight determination of pColA.
- Restriction endonuclease digestion (using HindIII, SmaI, AvaI, PstI, HincII) and analysis.
- Hybridization techniques using labeled colicin A mRNA.
Main Results:
- The ColA plasmid (pColA) was successfully amplified and purified.
- A molecular weight of 4.6 x 10^6 (7 kb) was determined for pColA.
- Twelve cleavage sites were mapped on pColA, ordered relative to a HindIII site.
- The structural gene for colicin A was localized to a 2.17-kb HincII fragment.
Conclusions:
- A detailed restriction map of the ColA plasmid (pColA) was established.
- The precise location of the colicin A structural gene was identified on the plasmid.