G.tigrina Hox gene DthoxC insertion into prokaryote E.coli / (by UNIamCloning)/2011/09/23

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Second DNA Extraction and Purification of the Three new samples of G.tigrina 9/23/11

  • Three more extractions were completed. Due to a poor DNA yield from the first extraction, it was decided to alter the amounts of planarian used. The first two samples (Gt. 2-1, Gt. 2-2) each contained half of one planarian. The third sample (Gt.3) contained two planarians. We were not sure if the low yield of DNA from the first extraction (Gt.1) was due to having too high of a concentration (too high of a concentration of DNA would overload the mini column in the kit producing an overall lower yield) or too low of a concentration of DNA which is why we halved our initial amount as well as doubled our initial amount of planaria. The DNA extraction was performed using the QIAGEN DNeasy Blood and Tissue Kit and the same procedure was used as the first extraction with a couple of minor changes. First, 2µl yeast tRNA was added along with the 200µl Buffer AL, mixed, and incubated at 56 degrees Celsius for ten minutes. The adding of the yeast tRNA will help with the amplification later during PCR. The incubation allowed for the activation of the yeast tRNA. Finally, the last change made during the DNA extraction was made after the addition of buffer AE. After the addition of the buffer AE, the sample was incubated at 65 degrees Celsius for five minutes.