Restriction enzymes products

After a desired clone is obtained and mapped with restriction enzymes, further analysis usually depends on the deterrnination of its nucleotide sequence. The nucleotide sequence of a new gene often provides clues to its function and the stmcture of the gene product. Additionally, the DNA sequence of a gene provides a guidepost for further manipulation of the sequence, for example, lea ding to the production of a recombinant protein in bacteria. 

Each of the -6000 PCR products was then co-transformed into yeast along with the recipient vector that had been linearized using a restriction enzyme that digests the plasmid at the desired cloning site. The 70 bp of homologous flanking sequence on each end of the PCR products is sufficient for the yeast homologous recombination system to act upon and insert the PCR product into the vector (Hudson et al., 1997 Ma et al., 1987). 

Enzyme modification, performance improvement, 3 671 Enzyme multiplied immunological technique (EMIT), 12 97 Enzyme Nomenclature, 17 402 Enzyme-product (EP) complex, 10 318 Enzyme production, Bacillus and, 12 477 Enzymes. See also Restriction enzymes Enzymes, 5 201... 

Biolabs (Pickering, ON), and Promega (Madison, WI). It is important to note that restriction enzymes producing 3 -overhangs should be avoided if possible (e.g., Psfl, Sfil, Kpnl). The use of such enzymes has been reported to result in the production of additional, nonspecific transcripts (Schenborn and Mierendorf, 1985). If these enzymes must be used, an exonuclease such as DNA Polymerase 1 Large (Klenow) Fragment can be utilized to convert the overhang to a blunt end before the template is transcribed. 

Flexible plate, 96 well, U-bottom without lid (BECTON DICKINSON, cat. No. 353911). There are several 96-well plates, but this product is easily handled especially for restriction enzyme digestion of a large number of DNA samples. 

Multiplex PCR amplification can apparently tolerate degraded DNA because the size of PCR products generated for subsequent hybridization is 100 50 base pairs. However, the 10 K mapping array and the 100 K and 500 K arrays use a specific restriction enzyme to digest the genomic DNA for subsequent ligation with a specific DNA linker. The DNA linkers then act as binding sites for the specific primer to initiate PCR amplification... 

Mix 5 -12 pi PCR product or restriction enzyme digest reaction mixture with... 

Restriction endonucleases are bacterial enzymes that cleave DNA at sequence specific sites. They were first discovered in 1970 [19]. Almost 2000 restriction enzymes have been identified since, and several hundred of these are commercially available [1]. Many mutations remove or create a particular restriction site in the DNA sequence. These mutations can be identified by PCR amplification, incubation of the product with the appropriate enzyme followed by visualisation of the fragments on an agarose gel. 

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