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Team:KAIST-Korea/Project/Methods
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To manipulate gene products obtained from gene-bank, our team used PCR/Real-Time PCR methods. Commercially obtained genes were containing few more base paires added at front and back site. As FGPR, the receptor protein of human, has human-specific signal peptide at its starting point, this region had to be replaced<br> | To manipulate gene products obtained from gene-bank, our team used PCR/Real-Time PCR methods. Commercially obtained genes were containing few more base paires added at front and back site. As FGPR, the receptor protein of human, has human-specific signal peptide at its starting point, this region had to be replaced<br> | ||
by ''S.pombe'' specific signal peptide to express the protein at membrane region. To do this, PCR primer containing signal peptide region of ''S.pombe'' were synthesized with method described below. Then, FGPR gene was amplifed by PCR to get gene product which its signal region is replaced. Also, additional sequence at rear site of gene was | by ''S.pombe'' specific signal peptide to express the protein at membrane region. To do this, PCR primer containing signal peptide region of ''S.pombe'' were synthesized with method described below. Then, FGPR gene was amplifed by PCR to get gene product which its signal region is replaced. Also, additional sequence at rear site of gene was | ||
| - | removed. STAT gene passed through | + | removed. STAT gene passed through similar process; removing additional region.<br> |
<br> | <br> | ||
PCR was done by commercial service sponsered by Bioneer.Inc (Korea), using AccuPower PCR system supplied by same company as reference below. | PCR was done by commercial service sponsered by Bioneer.Inc (Korea), using AccuPower PCR system supplied by same company as reference below. | ||
Revision as of 08:37, 22 July 2010
PCR
To manipulate gene products obtained from gene-bank, our team used PCR/Real-Time PCR methods. Commercially obtained genes were containing few more base paires added at front and back site. As FGPR, the receptor protein of human, has human-specific signal peptide at its starting point, this region had to be replaced
by S.pombe specific signal peptide to express the protein at membrane region. To do this, PCR primer containing signal peptide region of S.pombe were synthesized with method described below. Then, FGPR gene was amplifed by PCR to get gene product which its signal region is replaced. Also, additional sequence at rear site of gene was
removed. STAT gene passed through similar process; removing additional region.
PCR was done by commercial service sponsered by Bioneer.Inc (Korea), using AccuPower PCR system supplied by same company as reference below.
Oligo Synthesis
Homologous Recombination
References
Bioneer PCR service:
http://www.bioneer.co.kr/product/product_sub.jsp?pclassCode=1
Bioneer Oligo sythetis service
Put reference list here
