PCR - Bay/Sea Scallop DNA

An additional attempt to get the actin primers to work for use in screening samples for bay/sea scallop hybrids. The scallop_actin_fw primer was used in conjunction with the following:

bay_actin_Rv0 (Rxn 1)

bay_actin_Rv2 (Rxn 2)

sea_actin_Rv4 (Rxn 3)

sea_actin_Rv5 (Rxn 4)

PCR set up is here. Just used Bay or Sea scallop gDNA (chelexed). When/If get this working correctly, will start screening the hybrid samples. Anneal of 53C.

Lane 1 - 100bp Ladder

Lane 2 - Rxn 1: Bay

Lane 3 - Rxn 1: Sea

Lane 4 - Rxn 1: H2O

Lane 5 - Rxn 1: H2O

Lane 6 - Rxn 2: Bay

Lane 7 - Rxn 2: Sea

Lane 8 - Rxn 2: H2O

Lane 9 - Rxn 2: H2O

Lane 10 - Rxn 3: Bay

Lane 11 - Rxn 3: Sea

Lane 12 - Rxn 3: H2O

Lane 13 - Rxn 3: H2O

Lane 14 - Rxn 4: Bay

Lane 15 - Rxn 4: Sea

Lane 16 - Rxn 4: H2O

Lane 17 - Rxn 4: H2O

Lane 18 - 100bp Ladder

Results: Rxn 1 shows amplification with both Bay & Sea Scallop gDNA. The bands are close in size, but look like they would be more distinguishable if run on higher percentage gel and for a longer period of time to get better separation. However, there is contamination in one of the two water samples..

Rxn 2 shows amplification of only the Bay Scallop gDNA.

Rxn 3 shows amplification in both Bay & Sea Scallop gDNA and both bands are of the exact same size.

Rxn 4 shows no amplification in either set of gDNA.

Using the primers used in Rxn 1 will probably allows us to succesfully screen potential hybrids. Just need to remember to use high-percentage agarose gels and run samples for longer periods of time to get sufficient separation.