Molecular Cloning: A Laboratory Manual, Book 2 |
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Page 30
1 . Mix aliquots of the packaging mixture ( or concentrated bacteriophage
suspension ) containing 10 , 000 – 20 , 000 recombinant bacteriophages in a
volume of 50 ul or less with 0 . 2 ml of plating bacteria . Incubate for 20 minutes at
37°C . 2 .
1 . Mix aliquots of the packaging mixture ( or concentrated bacteriophage
suspension ) containing 10 , 000 – 20 , 000 recombinant bacteriophages in a
volume of 50 ul or less with 0 . 2 ml of plating bacteria . Incubate for 20 minutes at
37°C . 2 .
Page 40
Incubate the DNA solution prepared in step 2 for 5 minutes at 37°C . Transfer 2 .
5 ul of the solution to the first microfuge tube or well of the microtiter plate
containing the S1 reaction mixture . To the remainder of the DNA solution , add
150 units ...
Incubate the DNA solution prepared in step 2 for 5 minutes at 37°C . Transfer 2 .
5 ul of the solution to the first microfuge tube or well of the microtiter plate
containing the S1 reaction mixture . To the remainder of the DNA solution , add
150 units ...
Page 86
0 ) and mix . mix gently . Incubate 5 minutes at Incubate 15 minutes at Incubate 7
minutes at 20°C . 37°C . 20°C . Add 50 ul DMS stop solu - Add 240 ul hydrazine
Add 200 ul hydrazine tion ( at 0°C ) and mix . stop solution ( at 0°C ) stop solution
...
0 ) and mix . mix gently . Incubate 5 minutes at Incubate 15 minutes at Incubate 7
minutes at 20°C . 37°C . 20°C . Add 50 ul DMS stop solu - Add 240 ul hydrazine
Add 200 ul hydrazine tion ( at 0°C ) and mix . stop solution ( at 0°C ) stop solution
...
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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 |
Copyright | |
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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