Molecular Cloning: A Laboratory Manual, Book 2 |
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Page 24
This problem may be minimized by treating the cleaved vector with a
phosphatase before ligation to cDNA . If the double - stranded cDNA contains
one or more recognition sites for the restriction enzyme , it will be cleaved and
subsequently ...
This problem may be minimized by treating the cleaved vector with a
phosphatase before ligation to cDNA . If the double - stranded cDNA contains
one or more recognition sites for the restriction enzyme , it will be cleaved and
subsequently ...
Page 34
To generate deletion mutants , a recombinant plasmid ( or bacteriophage M13
replicative form DNA ) is linearized with a restriction enzyme that cleaves at one
end of the target sequence . The doublestranded linear DNA is digested with BAL
...
To generate deletion mutants , a recombinant plasmid ( or bacteriophage M13
replicative form DNA ) is linearized with a restriction enzyme that cleaves at one
end of the target sequence . The doublestranded linear DNA is digested with BAL
...
Page 27
Under these conditions , the nuclease cleaves essentially at random and
converts a small fraction of the DNA to linear molecules . Few of the molecules
are cleaved more than once by the enzyme . Digestion with a restriction enzyme
that ...
Under these conditions , the nuclease cleaves essentially at random and
converts a small fraction of the DNA to linear molecules . Few of the molecules
are cleaved more than once by the enzyme . Digestion with a restriction enzyme
that ...
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Contents
TEST LIGATION 3 29 | 3 |
SYNTHESIS OF THE FIRST STRAND OF cDNA 8 | 11 |
FRAGMENT OF E coli DNA POLYMERASE I 13 59 | 14 |
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Common terms and phrases
acid activity agarose aliquots allow amount amplification appropriate approximately bacteriophage bacteriophage T4 base buffer carried cDNA cells centrifugation Chapter chromatography cleaved clones colonies concentration constructed containing deletions denatured described determine digestion DNA polymerase dNTPs double-stranded efficiency electrophoresis et al ethanol expression Figure filters fragment gene genomic hybridization increase Incubate inserted interest isolated labeled length libraries ligation linear linkers method microfuge minutes mixture molecules mRNA mutagenesis mutagenic mutations Nucleic nucleotides obtained oligonucleotide original plaques plasmid plasmid DNA plate pool position possible prepared presence primer probe problem protein purified radiolabeled reaction recombinant region remove restriction enzyme room temperature samples screening sequence single single-stranded solution specific step Store strand synthesis target DNA target sequence template termini tion transfer transformants tube units usually vector volume