Molecular Cloning: A Laboratory Manual, Book 2 |
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Page 11-40
LABELING OF SYNTHETIC OLIGONUCLEOTIDES USING THE KLENOW
FRAGMENT OF E . coli DNA POLYMERASE I In some cases ( e . g . , when using
oligonucleotides as probes in Southern hybridizations ) , it is important to
radiolabel to ...
LABELING OF SYNTHETIC OLIGONUCLEOTIDES USING THE KLENOW
FRAGMENT OF E . coli DNA POLYMERASE I In some cases ( e . g . , when using
oligonucleotides as probes in Southern hybridizations ) , it is important to
radiolabel to ...
Page 13-8
However , even with these improvements , the Klenow enzyme does not routinely
yield as much sequence as the more processive Sequenase enzymes ( see
below ) . The enzyme will not efficiently copy homopolymeric tracts or other
regions ...
However , even with these improvements , the Klenow enzyme does not routinely
yield as much sequence as the more processive Sequenase enzymes ( see
below ) . The enzyme will not efficiently copy homopolymeric tracts or other
regions ...
Page 13-76
5 hours Titrate primer or use an alternative primer Repurify primer Titrate primer
Increase amount of Klenow fragment to 2 units per template Add additional
Klenow fragment during the chase ( see note to step 13 , page 13 . 63 ) Try a new
...
5 hours Titrate primer or use an alternative primer Repurify primer Titrate primer
Increase amount of Klenow fragment to 2 units per template Add additional
Klenow fragment during the chase ( see note to step 13 , page 13 . 63 ) Try a new
...
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Contents
CONCENTRATING NUCLEIC ACIDS E | 8-10 |
SYNTHESIS OF THE FIRST STRAND OF DNA 8 | 8-11 |
MEASUREMENT OF RADIOACTIVITY IN NUCLEIC ACIDS E | 8-18 |
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Common terms and phrases
aliquots allow amount amplification antibody appropriate approximately bacteriophage bacteriophage T4 base binding buffer carried cDNA cells centrifugation Chapter chromatography cleaved clones colonies complete concentration constructed containing deletions denatured described detection determine digestion dNTPs double-stranded efficiency electrophoresis et al ethanol expression extraction Figure filters four fragment gene genomic DNA hybridization increase Incubate inserted interest isolated Klenow labeled length libraries ligation linear linkers membrane method minutes mixture molecules mRNA mutants nucleic acids nucleotides obtained oligonucleotide plaques plasmid DNA plate position possible precipitation prepared presence primer probe problem protein purified radioactivity radiolabeled reaction recombinant region remove restriction enzyme room temperature samples screening separate sequence single single-stranded solution step stored strand synthesis target DNA target sequence template termini transfer Tris tube units usually vector volume