In today's lab we set up a PCR reaction that will be used to combine our two separate exons into one complete gene segment. Our two exons are ~800 BP and ~300 BP in length, and by using only our forward and reverse flanking primers we hope to end with one complete DNA fragment ~1100 BP in length. We chose to run our PCR at three different annealing temperatures (38, 40, and 42 degrees celcius), and also included a negative control for our DNA set (primers only,a positive control using a separate DNA sample, and a negative-positive control using different primers, but no DNA.
Two microliters of each exon DNA were used for each sample, and since this is highly concentrated with just the exon fragment, this amount should be sufficient for the PCR reaction. A gel electrophoresis will be run on Thursday to determine if our exons were successfully rejoined. We will not know for sure if this is our specific OOMT2 gene, but if the gene fragment is ~1100 BP, it will be the correct size, and then the sequence will be determined to see if we've successfully isolated our gene.
Written by Kim Lovik