Molecular Cloning: A Laboratory Manual, Book 2 |
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Results 1-3 of 13
Page 8-39
... ( HindIII / BamHI / EcoRI ) at codons 5-10 of the lacZ gene ( see map of λORF8 in Chapter 2 , page 2.50 ) . Insertion ... HindIII . Because M.HindIII methylase is not commercially available , the cDNA is methylated with M.BamHI methylase ...
... ( HindIII / BamHI / EcoRI ) at codons 5-10 of the lacZ gene ( see map of λORF8 in Chapter 2 , page 2.50 ) . Insertion ... HindIII . Because M.HindIII methylase is not commercially available , the cDNA is methylated with M.BamHI methylase ...
Page 8-40
... HindIII Agt18 , Agt19 encoded by the mcrB locus of E. Synthesize blunt - ended double - stranded cDNA Methylate with Alul and BamHI methylases Ligate to synthetic linkers BamHI GCTTGGATCCAAGCNNNNNNNN CGAACCTAGGTTCGNNNNNNNN linker AAAA ...
... HindIII Agt18 , Agt19 encoded by the mcrB locus of E. Synthesize blunt - ended double - stranded cDNA Methylate with Alul and BamHI methylases Ligate to synthetic linkers BamHI GCTTGGATCCAAGCNNNNNNNN CGAACCTAGGTTCGNNNNNNNN linker AAAA ...
Page 8-41
... HindIII site is formed by fusion of the poly ( dT : dA ) tract at the 3 ' terminus of the cDNA to the first linker ( see Figure 8.13 ) . Addition of linkers to the 5 ' terminus of the cDNA does not usually create a new restriction site ...
... HindIII site is formed by fusion of the poly ( dT : dA ) tract at the 3 ' terminus of the cDNA to the first linker ( see Figure 8.13 ) . Addition of linkers to the 5 ' terminus of the cDNA does not usually create a new restriction site ...
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acrylamide agarose gel aliquots amplification antibody bacteriophage M13 bacteriophage T4 bacteriophage T4 DNA buffer carried cDNA clones cDNA libraries cells centrifugation at 12,000g chromatography cleavage cleaved cleotide codon coli DNA polymerase complementary concentration containing denatured digestion dithiothreitol DNA ligase DNA sequencing DNAase dNTPs double-stranded DNA EDTA EDTA pH 8.0 efficiency ethanol exonuclease formamide gel electrophoresis gene genomic DNA H₂O hybridization Incubate the reaction Klenow fragment labeled ligation linear linkers method MgCl2 microfuge tube minutes at 4°C molecules mRNA mutagenesis mutations Natl nitrocellulose nitrocellulose filters Nucleic Acids nucleotides oligonucleotide phagemid plaques plasmid plasmid DNA plate pmoles polymerase chain reaction precipitation prepared primer probe purified radioactivity radiolabeled radiolabeled DNA recombinant remove restriction enzyme room temperature sample screening sequencing reactions single-stranded DNA solution specific activity stored strand of cDNA synthesis target DNA target sequence template DNA termini transcription transfer Tris Cl pH unlabeled vector