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Nucleic Acids Research, 1977, Vol. 4, No. 10 3455-3471
© 1977

Articles

The synthesis of high yields of full-length reverse transcripts of globin mRNA

E. Y. Friedman* and M. Rosbash**

Rosenstiel Basic Medical Sciences Research Center and Department of Biology, Brandeis University Waltham, MA 02154, USA

**To whom correspondence should be addressed

Received June 17, 1977.

Conditions have been determined under which reverse transcriptase catalyzes the synthesis of high yields of full length complementary deoxyribonucleic acid (cDNA). These conditions depend not only on the concentration of deoxynucleoside triphosphates (1) but also on the concentration of reverse transcriptase. An analysis of the kinetics of cDNA synthesis and the size of cDNA synthesized as a function of time under different conditions indicates that the mechanism of action of reverse transcriptase is partially distributive. This accounts for the neccessity of a high enzyme concentration to obtain high yields of full length cDNA. Additional experiments indicate that the yield of cDNA is limited by the fact that the template mRNA is rapidly inactivated. This is most likely due to the fact that the product cDNA is hydrogen bonded to the template mRNA during synthesis.

* Present address Department of Biochemistry, Johns Hopkins School of Hygiene and Public Health 615 North Wolfe Street Baltimore, Maryland 21205


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