Characterization of Plasmids prepared from Bacillus thuringiensis strains obtained from Middle Tennessee
Bacillus thuringiensis is a gram-positive, aerobic, spore-forming soil bacterium, which produces different insecticidal toxic proteins that are commercially used in controlling agriculturally important insects. B. thuringiensis strains are known to carry large plasmids on which genes for insecticidal proteins are often housed. Plasmids are self-replicating extra chromosomal DNA molecules found in Gram-negative and Gram-positive bacteria. The objectives of this study were to characterize strains of B. thuringiensis isolated from soil, dust from grain storage, and water samples collected in Middle Tennessee based on their plasmid content and to characterize the plasmids from these strains using restriction digestion analyses. In this research, plasmid DNA preparations were carried out on a total of 48 strains using QIAprep® Spin Miniprep Kit and analyzed by agarose gel electrophoresis in order to separate mixed DNA fragments. Agarose gel electrophoresis showed that all the B. thuringiensis strains contain at least one plasmid, and some strains contained as many as five (5) or more plasmids. In addition, three types of restriction enzymes (EcoRI, EvoRV, and HindIII) were used to digest the plasmids. The first enzyme EcoRI digested the plasmids of all the strains except B. thuringiensis18. The second enzyme was EcoRV, digested all of the plasmids of the B. thuringiensis strains except 13. Furthermore, HindIII digested the plasmids of all the strains except B.thuringiensis13, 14, 15, and 17. The plasmids were determined to range in size from 1,517 bp to 10,500 bp or larger. In conclusion, it was found that the Bt strains studied in this project were not unique in their plasmid content. Further studies should involve characterizing individual plasmids of these strains to determine the presence of unique genes. It would also be of interest to determine the presence of any restriction enzymes associated with these organisms.
Rasha Hadi Alsamadani,
"Characterization of Plasmids prepared from Bacillus thuringiensis strains obtained from Middle Tennessee"
ETD Collection for Tennessee State University.