Team:UEA-JIC Norwich/Weektwelve


University of East Anglia-JIC



Alistair performed PCR upon the CaMV promoter and ran on a gel which gave positive results showing a sequence on gels of about the right size in bp.


Activity resumes and Ben Jevans continuous his arduous routine of more PCR’s and gel running. This time it is with the arginine biosynthesis gene; results from the gel don’t tell anything too reassuring. Alistair performed PCR purification and then ran his CaMV promoter on a gel and nanodroped them. There was a significant decrease in DNA product concentration after PCR purification.


The day initiates with a nanodrop on all PCR products including the arginine biosynthesis gene, pSAD promoter, terminator and G-Luc gene. This was swiftly followed by DNA purification and then all components were ran on a gel were they showed a mixture of good and bad results.


Ben Jevans recoups the successfully purified products of the following the day and performs another nanodrop on them. Then a new technique to us known as poly ‘A’ tailing was executed upon the purified products to ensure greater stability. After this a vector ligation between the PCR products and the vector pGEM-T was performed, hoping all went according to plan. Alistair also performs the same experiment with pGEM-T using the CaMV promoter, but with no poly ‘A’ tailing as his sequence already had the A tail attached due to the polymerase he used.


A standard transformation was conducted using the newly ligated vector and then DNA purification was carried out on then pSAD terminator and arginine biosynthesis PCR products. This was also done with the CaMV promoter by Alistair. Alistair, Ben and Abbie gave a presentation to the JIC Metabolic Biology staff.


New cultures made in an LB broth containing ampicillin; these were made from colonies grown from previous cloning procedures and include the pSAD promoter, G-Luc and CaMV promoter.