Ran qPCRs on the O.lurida total RNA I isolated on 20150507 to assess presence of gDNA carryover with Oly Actin primers (SR IDs: 1505, 1504).

Used 1μL from all templates.

All samples were run in duplicate.

Positive control was HL1 O.lurida DNA isolated by Jake on 20150323.

Master mix calcs are here: 20150512_qPCR_Oly_RNA

Cycling params:

• 95C - 3mins

40 cycles of:

• 95C - 5s

• 60C - 20s

Melt curve

Plate layout: 20150512_qPCR_plate_Jake_Oly_Control_RNA

Results:

qPCR Data File (Opticon2): Sam_20150512_105811.tad

qPCR Report (Google Spreadsheet): 20150512_qPCR_Report_Jake_Oly_Control_RNA

Excluding the no template controls (NTC), all samples produced amplification. Will require DNasing before making cDNA.

On a side note, it should be noted that the efficiencies for all of the reactions were pretty bad; probably averaging 50%. Not entirely sure why or what that indicates.

In the amplification plots below, the positive control reps are the two red lines coming up at cycle ~22.

Amplification Plots

(http://eagle.fish.washington.edu/Arabidopsis/20150512_qPCR_Amp_Jake_Oly_Control_RNA.JPG)

Melt Curves

(http://eagle.fish.washington.edu/Arabidopsis/20150512_qPCR_Melt_Jake_Oly_Control_RNA.JPG)