| Budget Amount *help |
¥2,200,000 (Direct Cost: ¥2,200,000)
Fiscal Year 1991: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 1990: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 1989: ¥800,000 (Direct Cost: ¥800,000)
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| Research Abstract |
Restriction endonucleases have been screened in pathogenic Vibrios such as Vibrio choleras 01, Vibrio choleras non0l, Vibrio parahaemolyticus, Plesiomonas shigelloides and Aeromonas sobria, using lysozyme-lysis method described by us (Miyahara et al., Analyt. Chim. Acta, 213, 273-277, 1988). Restriction endonucleases were detected at a frequency of about 0.1 in these Vibrios. Among eleven restriction endonucleases purified and characterized, nine were isoschizomers of well-known restriction endonucleases, while the remaining two were new restriction endonucleases with a novel specificity. The designation of the two restriction endonucleases and the producer strains were PshAI in Plesiomonas shigelloides 319-73, and VpaK32I in Vibrio parahaemolyticus 4387-61. The former recognizes and cleaves the sequence 5'-GACNN/NNGTC, while the latter does the sequence 5'-GCTCTTC (1/4). Because of their stability and high yield, PshAI and VpaK32I should be useful for the recombinant DNA technology. It is of interest that all Vibrio parahaemolyticus restriction endonucleases found in this study recognized and cleaved non-palindromic or interrupted palindromic sequences, though the number of restriction endonucleases characterized was very limited. The result indicates that Vibrio parahaemolyticus may be a good source for restriction endonucleases recognizing non-palindromic sequences, as such restriction endonucleases have been found at very low frequencies in other genera of bacteria. The recognition specificities of Vibrio choleras non0l restriction endonucleases were GAATTC (EcoRI), AGGCCT (StuI), and AGTACT (ScaI).
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