Monday, August 10, 2015

New sequencing data.

Objective: The last PCR produced on july 22nd was prepared (ethanol precipitation) and sent out for Sanger sequencing. Today I wanted to look at the results and work with them to see if I could align.

Results: The results from the sequencing appear to be successful (shown below) . The HQ percentage is high (90.9, 82.7, 95.1, 96.3) and when aligned all four sequences align well between bases 80-120, 149-523, 569-624. I compared to previous sequences and showed some overlap with previous sequences. The red bars on each side of the sequences are areas that were trimmed due to low quality of the sequence. The top row is a consensus sequence created from the alignment of the for strands It shows the most likely bases when comparing all sequences to each other. With these sequences we should be able to create new primers that will allow me to qPCR my samples to check for gene expression moving forward with my research.

Quality and length of each strand

Alignment of all sequencing data (16 sequences) note: the new strands are the last four that show some overlap with the first 5 sequences.

Genius alignment of the four sequences

Reverse 1 sequnce
Reverse 2 sequence
 Forward 1 sequence
 Forward 2 sequence
 Consensous Sequence:

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