TH613A - Method for altering double stranded DNA - Google Patents

Abstract

This proposed invention addresses the problem of tryptophan repression and attenuation in a different manner and provides (1) a method to obtain an expression vector designed for the direct expression of an alien gene from the trp promoter-operator system; (2) a method to obtain a vector designed for the expression of a polypeptide that is specifically cut from the tryptophan operator-promoter and encoded by a gene obtained by linking the alien gene to the bacterial gene; and (3) a method of expression of the alien polypeptide that is well-controlled, highly efficient, and yields no less than related methods.

Claims (3)

1. The procedure for cutting double-stranded DNA at any given point consists of (a) converting the double-stranded DNA to single-stranded DNA in the region surrounding that point; (b) hybridizing it with the single-stranded region obtained in step (a) using an auxiliary primer of the same length as the single-stranded DNA, with the 5' end of the primer opposite to the nucleotides located 5' prior to the intended cutting point; (c) constructing the second segment eliminated in step (a), located in the 3' direction, from the aforementioned primer by reacting it with DNA polymerase containing deoxynucleotide triphosphate containing adenine, thymine, guanine, and cytosine; and (d) digesting the remaining length of single-stranded DNA that extends beyond the intended cutting point. 2. The procedure under claim 1 in which steps (c) and (d) occur concurrently by reaction with a DNA polymerase, which polymerizes in the 5'-3' direction, becoming exonucleolytic in the 3'-5' direction but non-exonucleolytic in the 5'-3' direction. 3. The process under claim 2, where the polymeric agent is Klenow Polymerase I (3 claims, 1 page, 13 figures).