Molecular Biology Plan for the Medicine and Biochemistry Society at NLCS Jeju
PCR
1. PCR a gene fragment using DNeasy PCR beads (premade dehydrated Taq + Buffers + dNTPs) from peanut DNA and then visualise it using agarose gel electrophoresis
2. Select a gene from an organism that has easily obtainable genomic DNA, create and order the necessary primers. Then extract DNA and then PCR but using a PCR reaction mix made from scratch by ourselves.
Gene cloning, cell culture and protein purification
3. Buy a transformation kit from Bio Rad (pGLO™ Bacterial Transformation Kit) website:
http://www.bio-rad.com/en-kr/product/pglo-bacterial-transformation-kit
Transform the bacteria (which means making competant E.coli cells take up a plasmid with a gene inserted into it for antibacterial resistance and for the green fluorescent protein(GFP).
4. Grow up a sample of GFP expressing bacterial cells, and the competant cells, add storage buffer (mostly glycerol) and freeze at -20 for later use.
5. Grow large amounts of transformed bacteria and try to separate the GFP using a simplified, homemade version of flow rate chromatography.
6. Run the protein extracts through a polyacrylamide electrophoresis gel.
Restriction digests and further PCR techniques
7. Grow up the bacterial cells, purify the plasmids and then do restriction digests on them.
8. Perform PCR on isolated target regions of either genomic or plasmid DNA within the Ecoli cells
Future work
9. Clone into the bacterial plasmids a protein of interest to confer upon the transformants a specific resistance to antibody or expressed protein leading to a novel ability that can be easily screened (e.g. UV resistance).
10. Investigate the lac operon, possibly using lac negative Ecoli strains. Possibly using the lac operon as a starting switch, e.g. to express GFP?
PCR
1. PCR a gene fragment using DNeasy PCR beads (premade dehydrated Taq + Buffers + dNTPs) from peanut DNA and then visualise it using agarose gel electrophoresis
2. Select a gene from an organism that has easily obtainable genomic DNA, create and order the necessary primers. Then extract DNA and then PCR but using a PCR reaction mix made from scratch by ourselves.
Gene cloning, cell culture and protein purification
3. Buy a transformation kit from Bio Rad (pGLO™ Bacterial Transformation Kit) website:
http://www.bio-rad.com/en-kr/product/pglo-bacterial-transformation-kit
Transform the bacteria (which means making competant E.coli cells take up a plasmid with a gene inserted into it for antibacterial resistance and for the green fluorescent protein(GFP).
4. Grow up a sample of GFP expressing bacterial cells, and the competant cells, add storage buffer (mostly glycerol) and freeze at -20 for later use.
5. Grow large amounts of transformed bacteria and try to separate the GFP using a simplified, homemade version of flow rate chromatography.
6. Run the protein extracts through a polyacrylamide electrophoresis gel.
Restriction digests and further PCR techniques
7. Grow up the bacterial cells, purify the plasmids and then do restriction digests on them.
8. Perform PCR on isolated target regions of either genomic or plasmid DNA within the Ecoli cells
Future work
9. Clone into the bacterial plasmids a protein of interest to confer upon the transformants a specific resistance to antibody or expressed protein leading to a novel ability that can be easily screened (e.g. UV resistance).
10. Investigate the lac operon, possibly using lac negative Ecoli strains. Possibly using the lac operon as a starting switch, e.g. to express GFP?