Week 5. Semester 3. Not good

Last Friday we re-ran a PCR for the stock positive controls and this Tuesday, September 29th, we ran a gel to analyze the banding. We were hoping the problem would be in the Primers we used last time and used brand newly aliquoted Primers but still got good bands. We pretty much had the same results are last time. There was a very small band at the appropriate distance for 16s (7/13/2015). Please refer to picture below.

Analysis on Agarose Gel Electrophoresis (9-29-15)

1% Agarose gel with 1μL/1% SYBER®Green.

Loaded:

Molecular Weight Marker (2 μL).

positive control DNA (2 μL)

loading dye (2 μL)

3 μL water

Loaded 7μL of control DNA sample and 2 μL of molecular weight marker.

Ran gel at 100 volts for 45-50 minutes in a 1x TAE Buffer solution (490mL dI water +10 mL 50xTAE)

Placed gel in gel doc to analyze banding.

 16s

Lane 1	Lane 2	Lane 3	Lane 4	Lane 5	Lane 6	Lane 7	Lane 8
	MWM		7-7-15		7-13-15	7-30-15

Week 4. Semester 3. Gel re-run and plans for today

This week we re-ran the gels for the previous unsuccessful PCR amplification of positive controls and got the same results. We used a freshly prepared buffer and new loading dye to make sure the problem was not in the gels. Today we will be working on re-running the PCR for the same positive controls with new primers in hopes we can get nice banding in the gels next week. Protocol:
Analysis on Agarose Gel Electrophoresis (9-22-15)

1% Agarose gel with 1μL/1% SYBER®Green.

Loaded:

Molecular Weight Marker (2 μL).

Positive control DNA (2 μL)

loading dye (2 μL)

3 μL water

Loaded the full 7 μL of control DNA mixture (PCR product, loading dye and water) and 2 μL of molecular weight marker.

Ran gel at 100 volts for 45-50 minutes in a 1x TAE Buffer solution (490mL dI water +10 mL 50xTAE)

Placed gel in gel doc to analyze banding.

16s 

Lane 1	Lane 2	Lane 3	Lane 4	Lane 5	Lane 6	Lane 7	Lane 8
MWM		7-7-15	7-8-15		7-13-15	7-30-15

mip

Lane 1	Lane 2	Lane 3	Lane 4	Lane 5	Lane 6	Lane 7	Lane 8
MWM		7-7-15	7-8-15		7-13-15	7-27-15