منابع مشابه
PCR-mediated Expression of the Human GM-CSF Gene in Escherichia coli
Four exons of the human genomic GM-CSF gene were assembled together using gene splicing by overlap extension (SOE) method. The resulting nucleotide sequence was cloned in the pET23a(+) expression vector under the control of strong bacteriophage T7 transcription and translation signals. The construct obtained was Transferred into the E. coli strain, BL21(DE3) pLysS and IPTG was used for inducti...
متن کاملRT-PCR MEDIATED CLONING OF HUMAN GROWTH HORMONE GENE AND I TS EXPRESSION IN E. coli
Human growth hormone (hGH) genomic sequence containing 5 exons and 4 introns was cloned in pcDNA-3 and the constructed plasmid was subsequently used for transfection ofNlli-3T3 cell line using lipofection technique. Expression of hGH in stably transfected cells was assayed using ELISA. Total RNA was extracted from transfected cells and hGH cDNA was amplified by RT-PCR using specific primers...
متن کاملPCR-mediated direct gene disruption in Schizosaccharomyces pombe.
We have examined the feasibility and efficiency of PCR-mediated direct gene disruptions in the fission yeast Schizosaccharomyces pombe. In the present study, the S.pombe ura4+ gene was amplified by PCR with oligonucleotides that had short flanking regions ( approximately 40 bp) to the target gene. Using this purified PCR product we were able to disrupt genes in an S. pombe strain bearing aura4 ...
متن کاملPCR-based gene synthesis to produce recombinant proteins for crystallization
BACKGROUND Gene synthesis technologies are an important tool for structural biology projects, allowing increased protein expression through codon optimization and facilitating sequence alterations. Existing methods, however, can be complex and not always reproducible, prompting researchers to use commercial suppliers rather than synthesize genes themselves. RESULTS A PCR-based gene synthesis ...
متن کاملPCR- and ligation-mediated synthesis of marker cassettes with long flanking homology regions for gene disruption in Saccharomyces cerevisiae.
We developed a novel method for synthesizing marker-disrupted alleles of yeast genes. The first step is PCR amplification of two sequences located upstream and downstream of the reading frame to be disrupted. Due to the addition of non-specific single A overhangs by Taq DNA polymerase, each PCR product can be ligated with a marker DNA which has T residues at its 3' ends. After amplification of ...
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ژورنال
عنوان ژورنال: Nucleic Acids Research
سال: 1989
ISSN: 0305-1048,1362-4962
DOI: 10.1093/nar/17.11.4403