Application of partial restriction procedure in both shotgun and non-random strategies for nucleotide sequencing |
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Authors: | Bing Zhou Nongan Chen and Qiliang Li |
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Institution: | Shanghai Institute of Biochemistry, Chinese Academy of Sciences, Shanghai 200031, China |
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Abstract: | Partial digestion of a target DNA fragment with 4-bp-recognition restriction enzymes followed by a forced ligation to an M13 vector was employed for the construction of a subfragment library. The library can be used for either shotgun or non-random nucleotide sequencing. Application of the partial digests generated with the 4-bp recognition restriction enzymes instead of DNase I in the improved non-random strategy for nucleotide sequencing (Li and Wu, 1987) made the procedure as easy as that of the random strategy. The library can also be used in shotgun nucleotide sequencing directly, and few self-ligated subfragments were found. The usefulness of this procedure was demonstrated by the sequencing of a goat 6.5-kb EcoRI fragment, which is located 5' to the globin gene. |
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Keywords: | Recombinant DNA phage M13 vector restriction enzymes globin gene region goat |
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