Serrano, E. and Carrasco, B. (2019) Measurement of the length of the Integrated donor DNA during Bacillus subtilis natural chromosomal transformation. Bio-Protocol, 9(16), e3338. (doi: 10.21769/BioProtoc.3338) (PMID:33654843) (PMCID:PMC7854159)
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Abstract
For natural transformation to occur, bacterial cells must first develop a programmed physiological state called competence. Competence in Bacillus subtilis, which occurs only in a fraction of cells, is a transient stress response that allows cells to take up DNA from the environment. During natural chromosomal transformation, the internalized linear single-stranded (ss) DNA recombines with the identical (homologous) or partially identical (homeologous) sequence of the resident duplex. The length of the integrated DNA, which can be measured, depends on the percentage of sequence divergence between the donor (internalized) and the recipient (chromosomal) DNAs. The following protocol describes how to induce the development of competence in B. subtilis cells, how to transform them with donor DNAs representing different percentages of sequence divergence compared with the recipient chromosomal DNA, how to calculate the chromosomal transformation efficiency for each of them, and how to amplify the chromosomal DNA from the transformants in order to measure the length in base pairs (bp) of the integrated donor DNA.
| Item Type: | Articles |
|---|---|
| Status: | Published |
| Refereed: | Yes |
| Glasgow Author(s) Enlighten ID: | Serrano, Dr Ester |
| Authors: | Serrano, E., and Carrasco, B. |
| College/School: | College of Medical Veterinary and Life Sciences > School of Infection & Immunity |
| Journal Name: | Bio-Protocol |
| Publisher: | Bio-Protocol LLC |
| ISSN: | 2331-8325 |
| ISSN (Online): | 2331-8325 |
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