Expression of enzymatically active reverse transcriptase

C - Chemistry – Metallurgy – 12 – N

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195/1.18, 195/1.

C12N 1/21 (2006.01) C07K 14/15 (2006.01) C12N 9/12 (2006.01) C12N 15/10 (2006.01) C12N 15/54 (2006.01)

Patent

CA 1314504

EXPRESSION OF ENZYMATICALLY ACTIVE REVERSE TRANSCRIPTASE Abstract of The Disclosure This invention provides a plasmid which, when intro- duced into a suitable host cell and grown under appro- priate conditions, effects expression of a gene on the plasmid and production of a polypeptide having reverse transcriptase activity. The plasmid is a double- stranded DNA molecule which includes in a 5' to 3' order the following: a DNA sequence which includes an inducible promoters a DNA sequence which includes an ATG initiation codon; the central portion of the Moloney murine leukemia virus (MuLV) pol gene, said central portion including a DNA sequence which encodes the polypeptide having reverse transcriptase activity; DNA sequence which contains a gene associated with a selectable or identifiable phenotypic trait which is manifested when the vector is present in the host cell; and a DNA sequence which contains an origin of replica- tion from a bacterial plasmid capable of autonomous replication in the host cell. The invention also concerns a method for recovering purified enzymatically-active polypeptide having re- verse transcriptase activity, the polypeptide being encoded by the plasmid pB6 815.23, from a suitable host cell e.g. E. coli HB101 producing the polypeptide. finally, the invention concerns use of the polypeptide to prepare complementary DNA (cDNA).

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