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Abstract
The plasmid pBR322 had been observed to be excluded from bacteria also containing pUC19. To investigate whether plasmid size and the presence of the rop gene are involved in the exclusion of pBR322 by its successor pUC19, removal of a 1.1kb fragment from the pBR322 vector was attempted to produce pBR322 ∆ (1.1 rop ). This alteration reduced the size of pBR322 by 26%, making it closer in size to pUC19. It also removed the rop gene, the product of which has been implicated in regulation of DNA replication. Initial preparation of pBR322 and pUC19 DNA from HB101 E.coli was successful. Removal of the 1.1 kb fragment with Afl III and Sty I appeared to be successful, however subsequent ligation and transformation of DH5 α E.coli with pBR322 ∆ (1.1 rop ) proved to be challenging. Competent cells were successfully generated and DNA transformation was successful. After restriction digest analysis, however, clones isolated appeared to contain both pBR322 and pBR322 ∆ (1.1 rop ). Restriction enzymes employed in this analysis and technical problems with gel electrophoresis posed challenges within the timeline of the study. Thus it is not clear from this work how the reduction in size of pBR322 or the elimination of rop expression would affect the exclusion of pBR322 by pUC19.