Molecular Cloning: A Laboratory Manual, Book 2 |
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Page 8
Sedimentation through sucrose gradients formed in nondenaturing solvents
results in good recovery , but the presence of secondary structure in the RNA
often confounds effective fractionation . The method of choice , therefore , is
sucrose ...
Sedimentation through sucrose gradients formed in nondenaturing solvents
results in good recovery , but the presence of secondary structure in the RNA
often confounds effective fractionation . The method of choice , therefore , is
sucrose ...
Page 22
The ligation step is less important if the DNA is fragmented by digestion with
DNAase I in the presence of Mn * * . However , it is still worthwhile because it
helps to ensure that all segments of the target DNA will be equally represented in
the ...
The ligation step is less important if the DNA is fragmented by digestion with
DNAase I in the presence of Mn * * . However , it is still worthwhile because it
helps to ensure that all segments of the target DNA will be equally represented in
the ...
Page 27
CLEAVAGE OF DOUBLE - STRANDED CLOSED CIRCULAR DNA WITH
PANCREATIC DNAase I IN THE PRESENCE OF Mn + + The following protocol is
an adaptation of methods described by Anderson ( 1981 ) and Labeit et al . (
1987 ) .
CLEAVAGE OF DOUBLE - STRANDED CLOSED CIRCULAR DNA WITH
PANCREATIC DNAase I IN THE PRESENCE OF Mn + + The following protocol is
an adaptation of methods described by Anderson ( 1981 ) and Labeit et al . (
1987 ) .
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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