Transformasi Fragmen DNA Salmonella Ke Dalam Sel E. coli DH5-α

https://doi.org/10.22146/agritech.19320

E.S. Rahayu(1*), A. Pertiwiningrum(2), R. Indrati(3), S. Raharjo(4), S. Rahayu(5), S. Margino(6)

(1) Fakultas Teknologi Pertanian, Universitas Gadjah Mada
(2) Fakultas Peternakan, Universitas Gadjah Mada
(3) Fakultas Teknologi Pertanian, Universitas Gadjah Mada
(4) Fakultas Teknologi Pertanian, Universitas Gadjah Mada
(5) BPTP Ungaran, Jawa Tengah
(6) Fakultas Pertanian, Universitas Gadjah Mada
(*) Corresponding Author

Abstract


Genomic DNA from Salmonella typhimarium ATCC 14028, cultured in a Luria broth for 24 hr at 37°C, was isolated from the cells using EDTA as a chelating agent and SDS as a detergent and lysing agent. The amount of genomic DNA was then digested with each of three restriction endonucleases, i.e., EcoRI, HindlII, and BamHI for 4 hr at 37°C. A plasmid (pUC19) was used as a vector. Prior to use, the pUC19 plasmid was split with one of the three restriction enzymes and dephosphorylated using a bacterial alkaline phosphatase. The genomic DNA was then ligated to the corresponding prediggested plasmid using a ligase at 15°C over night. Transformation of the DNA recombinant into E.coli DH5-a was successfully carried out using a heat shock method as indicated by gel electrophoresis

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DOI: https://doi.org/10.22146/agritech.19320

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