PCR - (Oct/01/2012 )

Hi there,

I'm an agriculture undergraduate in 3rd yr and just completed the a DNA extraction on about 60 samples using a Qiagen flexigene DNA kit and looking to get some tips for the next phase which is PCR. Basically I am looking for the Booroola gene in the samples.

I've got my two primers for the gene and the redtaq polymerase...the university has a thermocycler and the agarose gel equipment.

Below (or attached) is an excel file with the calculations my supevisor has provided me on one sheet, the other sheet is the results of the UV absorbance using a Cecil spectrophotometer which shows the purity of the DNA samples, which 1.8 is the goal though as I'm only after 140kb then I should have plenty of DNA based on the UV 260nm specs (see Table 5a in the excel file sheet titled 'Master mix calculations').

What is the question exactly?

Hi Curtis,

Sorry I was a bit tired when I posted this and had edited my question before posting. Do you feel like critiquing the data in the last 2 sheets? Is there any other calculations I should be aware of for PCR?

Not particularly confident of the next steps, so difficult to know what data can be compiled from a PCR?