Tag Archives: 15:09-184

DNA Quantification – Ava’s RLO Trasmission Samples

Quantified DNA extractions from Ava’s samples that I isolated earlier this month, as well as some older samples that I hadn’t quantified yet.

Used the Roberts Lab Qubit 3.0 and the Qubit hsDNA Kit (high sensitivity). Used 5uL of template for all samples.

All data was added to the master extraction spreadsheet (Google Sheet): ava_abalone_master_extraction_list

Results:

20171026_Ava_RLO_quantification_qubit_01 (Google Sheet)

20171026_Ava_RLO_quantification_qubit_02 (Google Sheet)

There were 65 samples with concentrations that were too high for the high sensitivity assay. Will re-quantify this samples using the broad range assay.

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qPCR – Ava’s RLO Transmission Samples

Ran five plates of qPCRs on bunch of DNA I extracted ealier this month.

Standard curve was p18RK7 from 20161128.

All samples were run in duplicate.

Master mix calcs are here (Google Sheet): 20171026_qPCR_WSN1_Ava_Samples

Plate layouts, cycling params, etc. can be seen in the corresponding qPCR Reports (see Results below).

Baseline threshold was manually set to 580, based on Lisa’s development of the withering syndrome qPCR assay.

Results:

Curves looked great on each plate. See images at bottom of post.

NOTE: Plate #5 had a missed rep; I think this was due to a bad pipette tip. I excluded the 3e4 point of the curve in the analysis.

PLATE #1

qPCR Report (PDF): Sam_2017-10-26 07-31-12_CC009827.pdf
qPCR Data File (CFX96): Sam_2017-10-26 07-31-12_CC009827.pcrd

PLATE #2

qPCR Report (PDF): Sam_2017-10-26 09-02-31_CC009827.pdf
qPCR Data File (CFX96): Sam_2017-10-26 09-02-31_CC009827.pcrd

PLATE #3

qPCR Report (PDF): Sam_2017-10-26 10-32-10_CC009827.pdf
qPCR Data File (CFX96): Sam_2017-10-26 10-32-10_CC009827.pcrd

PLATE #4

qPCR Report (PDF): Sam_2017-10-26 12-01-33_CC009827.pdf
qPCR Data File (CFX96): Sam_2017-10-26 12-01-33_CC009827.pcrd

PLATE #5

qPCR Report (PDF): Sam_2017-10-26 13-31-05_CC009827.pdf
qPCR Data File (CFX96): Sam_2017-10-26 13-31-05_CC009827.pcrd


PLATE #1


PLATE #2


PLATE #3


PLATE #4


PLATE #5

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DNA Isolation – Ava Withering Syndrome Transmission Study Tissues

Isolated DNA from 144 red abalone digestive gland tissue samples.

Tissue was weighed, minced with a razor blade, and transferred to 2mL snap cap tube containing 1mL of InhibtEX Buffer.

DNA was extracted using the QIAmp Fast DNA Stool Mini Kit (Qiagen) following the manufacturer’s protocol with the following options:

Minced tissue was incubated at 70C O/N
Followed “human DNA analysis” protocol (to maximize sample recovery)
Eluted DNA with 100μL Buffer ATE
Sample information is in this spreadsheet (Google Sheet): ava_abalone_master_extraction_list

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