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Nucleic Acids Research, 1982, Vol. 10, No. 18 5429-5445
© 1982

MOLECULAR BIOLOGY

Accurate in vitro splicing of human ß-globin RNA

Ryszard Kole and Sherman M. Weissman

Department of Human Genetics, Yale University School of Medicine New Haven, CT 06510, USA

Received June 21, 1982. Revised August 19, 1982. Accepted August 19, 1982.

Human ß-globin RNA transcribed from an exogenous DNA template is spliced in vitro by concentrated whole cell extracts from HeLa cells. Using the primer extension technique, we have shown that the small intervening sequence is spliced accurately and that the sequence of the product across the splice junction 1s identical to that of ß-globin mRNA prepared from human reticulocytes.

The efficiency of the splicing reaction is low. The RNA transcript containing both introns and terminated upstream from the polyadenylation site is spliced most efficiently. The transcript which is terminated downstream from the polyadenylation site is not spliced at all.

Thalassemic ß-globin RNA which carries an extra splice site 1n the small intron is also spliced, albeit with a low yield.


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