Molecular Cloning: A Laboratory Manual, Book 2 |
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Page 8-24
... ligation of two unrelated cDNAs . The ligation mixture should therefore always contain a high molar excess of vector DNA to cDNA . However , these conditions strongly favor re - formation of the vector by self - ligation , leading to ...
... ligation of two unrelated cDNAs . The ligation mixture should therefore always contain a high molar excess of vector DNA to cDNA . However , these conditions strongly favor re - formation of the vector by self - ligation , leading to ...
Page 13-24
... Ligation of the Target DNA 1. Using the appropriate restriction enzymes , digest a sufficient amount of recombinant ... ligation conditions as described in the note to step 2. If it is not possible to avoid restriction enzymes that ...
... Ligation of the Target DNA 1. Using the appropriate restriction enzymes , digest a sufficient amount of recombinant ... ligation conditions as described in the note to step 2. If it is not possible to avoid restriction enzymes that ...
Page 15-10
... ligation buffer amount calculated above 100 ng 2 μ . blunt - end ligation buffer should be added last to reduce the possibility that the DNA will be precipitated by spermidine . Add 1-2 Weiss units of bacteriophage T4 DNA ligase , and ...
... ligation buffer amount calculated above 100 ng 2 μ . blunt - end ligation buffer should be added last to reduce the possibility that the DNA will be precipitated by spermidine . Add 1-2 Weiss units of bacteriophage T4 DNA ligase , and ...
Contents
E | 8-8 |
SYNTHESIS OF THE FIRST STRAND OF CDNA 8 | 8-11 |
CONCENTRATING NUCLEIC ACIDS BY EXTRACTION WITH BUTANOL E | 8-16 |
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acrylamide agarose gel aliquots amplification annealing antibody bacteriophage M13 bacteriophage T4 bacteriophage T4 DNA buffer carried cDNA clones cDNA libraries cells centrifugation at 12,000g cleavage cleaved cleotide coli DNA polymerase concentration containing ddNTPs deletions denatured digestion diluted dithiothreitol DNA ligase DNA sequencing DNAase dNTPs double-stranded DNA EDTA EDTA pH 8.0 ethanol exonuclease expression libraries formamide fusion proteins gel electrophoresis gene genomic DNA H₂O hybridization Klenow fragment labeled ligation linear linkers Maxam-Gilbert method MgCl2 microfuge tube minutes at 4°C molecules mRNA Natl nitrocellulose nitrocellulose filters Nucleic Acids nucleotides oligonucleotide phagemid piperidine plaques plasmid plasmid DNA plate polymerase chain reaction precipitation prepared primer probe purified radioactivity radiolabeled recombinant remove restriction enzyme room temperature samples Sequenase sequencing gel sequencing reactions sonicated specific activity stored strand of cDNA synthesis T4 DNA polymerase target DNA target sequence template DNA termini transcription transfer Tris Cl pH vectors µg/ml