Tag Archives: UW06:06-50

Data Summary – Black Abalone Phage qPCRs

A quick summary table of the various black abalone qPCRs I ran yesterday:

SAMPLE RLO_MCP RLO_ph_protease XC_prophage_portal RLOv_DNA_helicase WSN
06:06-50  +  +  +  +  +
06:06-52  +  +  +  +  +
07:12-01  -  -  -  +  -
07:12-02  -  -  -  -  -
08:13-05  +  +  +  -  +
08:13-18  +  +  +  -  +
08:13-24  +  +  +  -  +*
08:13-25  +  +  +  -  +
  • This sample technically showed amplification, but came up after the last point on the standard curve. Most likely due to extremely low concentration (~0.5ng/uL).

  • RLO Major Capsid Protein (RLO_MCP)

  • RLO Prohead Protease Protein (RLO_ph_protease)
  • XenoCal Phage Portal Gene (XC prophage)
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qPCR – WSN on Black Abalone

Ran qPCRs on a set of black abalone digestive gland DNA (sample list provided by Carolyn):

07:12-01 (Black Ab exp 1)
07:12-02 (Black Ab exp 1)
08:13-05 (Black Ab exp 2)
08:13-18 (Black Ab exp 2)
08:13-24 (Black Ab exp 2)
08:13-25 (Black Ab exp 2)
UW06:06-32
UW06:06-41
UW06:06-50 (Black Ab exp 1)
UW06:06-52 (Black Ab exp 1)

The two samples with a strikethrough did not have any DNA left in the tubes and were not run.

All samples were run in duplicate.

Standard curve was p18RK7 from 20161128.

Cycling params, plate layout, etc can be seen in the qPCR Report (see Results).

Baseline was set 580 as previously determined by Lisa.

Results:
qPCR Report (PDF): Sam_2017-04-13%2016-20-54_CC009827_WSN1.pdf
qPCR Data File (CFX): Sam_2017-04-13%2016-20-54_CC009827_WSN1.pcrd

Standard curve looked good.

The following samples did not amplify:
– 07:12 set
– Note: 08:13-24 technically did amplify, but comes up below the lowest point of the standard curve, so technically it is effectively “no amplification”.

The remaining samples all came up positive.

Will convey to Carolyn and Stan.

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qPCR – RLOv DNA Helicase on Black Abalone

Ran qPCRs on a set of black abalone digestive gland DNA (sample list provided by Carolyn):

07:12-01 (Black Ab exp 1)
07:12-02 (Black Ab exp 1)
08:13-05 (Black Ab exp 2)
08:13-18 (Black Ab exp 2)
08:13-24 (Black Ab exp 2)
08:13-25 (Black Ab exp 2)
UW06:06-32
UW06:06-41

UW06:06-50 (Black Ab exp 1)
UW06:06-52 (Black Ab exp 1)

The two samples with a strikethrough did not have any DNA left in the tubes and were not run.

All samples were run in duplicate.

Standard curve was from 20161106.

Cycling params, plate layout, etc can be seen in the qPCR Report (see Results).

Baseline was set 580.5 as previously determined.

Results:
qPCR Report (PDF): Sam_2017-04-13%2016-20-54_CC009827_RLOv_helicase.pdf
qPCR Data File (CFX): Sam_2017-04-13%2016-20-54_CC009827_RLOv_helicase.pcrd

Standard curve looked good, although efficiency is pushing it on the high end.

The following samples did <em>not</em> amplify:

  • 07:12-02
  • All 08 samples.

The remaining samples all came up positive, with the 06 set being extremely hot (came up around cycle 13).

Will convey to Carolyn and Stan.

 

 

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qPCR – RLO Prophage Genes

Ran qPCRs on a set of black abalone digestive gland DNA (sample list provided by Carolyn):

07:12-01 (Black Ab exp 1)
07:12-02 (Black Ab exp 1)
08:13-05 (Black Ab exp 2)
08:13-18 (Black Ab exp 2)
08:13-24 (Black Ab exp 2)
08:13-25 (Black Ab exp 2)
UW06:06-32
UW06:06-41

UW06:06-50 (Black Ab exp 1)
UW06:06-52 (Black Ab exp 1)

The two samples with a strikethrough did not have any DNA left in the tubes and were not run.

Gene targets:
– RLO Major Capsid Protein (RLO_MCP)
– RLO Prohead Protease Protein (RLO_ph_protease)
– XenoCal Phage Portal Gene (XC prophage)

Master mix calcs are here (Google Sheet): 20170413 – qPCR_XCphagePortal_RLOcapsid_RLOprohead

The same master mix calculations were used for each, just swapped in appropriate primers.

All samples were run in duplicate.

Cycling params, plate layout, etc. can be found in the qPCR Report (see Results below).

Results:
qPCR Report (PDF): Sam_2017-04-13 14-56-03_CC009827.pdf
qPCR Data File (CFX): Sam_2017-04-13 14-56-03_CC009827.pcrd

Melt curves for all three primer sets looked perfect (see below)

Amplification present for all samples, with all three primers except the 07:12 samples.

Will pass info along to Carolyn and Stan.

Will add info to the following two spreadsheets (Google Sheets):

Black Abalone: Expt 1 – WS & Phage

Black Abalone: Expt 2 – WS only

 


 

Green = RLO_ph_protease

Brown = RLO_MCP

Blue = XC_prophage

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