Vectors for in vitro production of RNA copies of either strand o

Details Vectors for in vitro production of RNA copies of either strand o Vectors for in vitro production of RNA copies of either strand o

1985-07-17

1988-08-23

Wiseman, Thomas G.

Chemistry: molecular biology and microbiology

Micro-organism, tissue cell culture or enzyme using process...

Preparing compound containing saccharide radical

4351723, 435235, 435320, 935 3, 935 16, 935 17, 935 29, 935 41, C12P 1934, C12N 1500, C12N 700

Patent

active

047660724

ABSTRACT:
In vitro production of RNA copies of either strand of any cloned DNA sequence may be obtained utilizing a unique cloning vector having two different opposed promoter sequences which are separated by a multiple cloning site. RNA polymerases which recognize only one of the particular promoter sequences in the vector may be applied to obtain transcription which proceeds from the recognized promoter toward the other promoter. Transcription of a desired strand of any DNA sequence is obtained by cleaving a particular restriction site in the multiple cloning site between the two promoter sequences, cloning the desired DNA sequence into the cleaved site, then cleaving another site between the two promoters which is distal to the promoter from which transcription is desired. The RNA polymerase which recognizes the selected promoter may then be applied to the vector to obtain transcription of the selected DNA sequence in vitro. Double stranded RNA may also be formed utilizing the vector by providing multiple vectors cleaved on either side of the DNA segment and thereafter applying the two RNA polymerases to cause transcription of both strands of the selected DNA segment. Specific cloning vectors are novel plasmids designated pGEM-1 and pGEM-2 which are characterized by having the SP6 and T7 late phage promoters facing each other separated by a multiple cloning site.

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