Using Geneious, we were able to match our fish DNA to DNA in the database. To do this we first copied the reverse sequence from the ‘Fish barcode Reverse Reads’ folder and pasted it into the ‘Forward Reads’ folder. Then, we selected the forward and reverse sequences of the same ID and clicked the Align/Assemble tab, choosing ‘De novo assembly”. Using the default settings, a new file appeared containing the consensus sequence for that fish DNA. We edited the sequence to clean up any discrepancies and saved these changes. We then right-clicked this file and chose ‘Generate consensus sequence’. We used this file to BLAST our sequence by right-clicking the file and selecting ‘BLAST’, keeping the default settings. Another file appeared, allowing us to see a set of top matches for our sequence. These matches told us what species our fish DNA most closely resembles. We created a new folder for a fish barcode test alignment and selected the assembly consensus sequence and 5-10 hits from the BLAST search, pasting these into the new folder. We then selected all of these sequences in the new folder and right-clicked, choosing ‘Multiple Align’ and then ‘Muscle Alignment’ with default settings. A new file was generated, showing polymorphisms between our DNA and other DNA of the same species.
Unfortunately, only two of my DNA barcodes were successful: KRS1 and KRS4.
|Number||Unique ID code||Restaurant Species Name||DNA Barcode Species Name|
|1||KRS1||Tuna||Thunnus Albacares (Yellowfin Tuna)|
|4||KRS4||Salmon||Salmon Salar (Atlantic Salmon)|
The successful DNA barcodes matched with the species specified at the restaurant.
For KRS1, there were 11 polymorphic sites found in columns 31, 167, 228, 324, 332, 333, 344, 346, 371, and 387.
For KRS4, there were 10 polymorphic sites found in columns 69, 586, 590, 611, 612, 614, 625, 628, 631, and 632.