Tag Archives: Opticon2

qPCRs – Tim’s Adult Gigas gill cDNA (from 20091009)

Duplicates of earlier qPCRs.

Primers: Cg_HIF1 and IL17 Iso D.

The previous version of IL17 primers used (IL17 internal) were NOT the ones used for the paper. The IL17 Iso D are the correct primers and are the same ones that Tim previously used on the juvenille gill samples.

qPCR set up and plate layout can be found here.

Results:

Duplicates of earlier qPCRs.

Primers: EF1 and IL17 Iso D.

The previous version of IL17 primers used (IL17 internal) were NOT the ones used for the paper. The IL17 Iso D are the correct primers and are the same ones that Tim previously used on the juvenille gill samples. qPCR set up and plate layout can be found here.

Results:

qPCR – BB & DH cDNA (from 20091223)

qPCR was set up on these cDNAs using the following primers:

FP008495.p.cg.6 (“GSTO1″, “Glutathione S-transferase omega-1″) – This was upregulated in BB SOLiD data.

These were run in duplicate to take up a full PCR plate. qPCR set up and plate layout can be found here.

Results:

qPCR Data File (Opticon): 20091231_152520.tad

Data workup: PROPS BB_DH Gene Expression Miner Workup

qPCRs – BB & DH cDNA (from 20091223)

qPCR was set up on these cDNAs using the following primers:

AJ582629.p.cg.6 (“DEF1″, “Defensin 1″) – This was upregulated in BB SOLiD data.

CB617519.p.cg.6 (“RETST”, “All-trans retinol”) – This was upregulated in BB SOLiD data.

qPCR set up and plate layout can be found here.

Results:

qPCR Data File (Opticon): 20091230_173747.tad

Data workup: PROPS BB_DH Gene Expression Miner Workup

 

 

qPCR was set up on these cDNAs using the following primers:

CU684779.p.cg.6 (“SEMSA”, “Semaphorin-SA”) – This was upregulated in BB SOLiD data.

FP004879.p.cg.6 (“TIMP3″, “Metalloproteinase inhibitor 3″) – This was upregulated in BB SOLiD data.

qPCR set up and plate layout can be found here.

Results:

qPCR Data File (Opticon): 20091230_143643.tad

Data workup: PROPS BB_DH Gene Expression Miner Workup

qPCR – BB & DH cDNA (from 20091223)

qPCR was set up on these cDNAs using the following primers:

FP010108.p.cg.6 (“DJB12″, “DnaJ homolog subfamily B member 12″) – This was upregulated in DH SOLiD data.

AJ565670.p.cg.6 (“TOP1″, “DNA topoisomerase 1″) – This was upregulated in BB SOLiD data.

qPCR set up and plate layout can be found here.

Results:

qPCR Data File (Opticon): 20091229_164912.tad

Data workup: PROPS BB_DH Gene Expression Miner Workup

qPCR – BB & DH cDNA (from 20091223) and Emma primer sets for testing

qPCR was set up on these cDNAs using the following primers:

EW778389.p.cg.6 (“DPGN”, “Serine protease inhibitor dipetalogastin”) – This was upregulated in DH SOLiD data.

FP001672.p.cg.6 (“PGSC1″, “Peptidoglycan-recognition protein SC1a/b”) – This was upregulated in DH SOLiD data.

Included three primer sets of Emma’s (matrillin, beta tub and chaperonin). These were set up with no template and done in duplicate.

qPCR set up and plate layout can be found here.

Results:

qPCR Data File (Opticon): 20091229_133148.tad

Data workup: PROPS BB_DH Gene Expression Miner Workup

Emma’s “beta tub” primers show some weird fluorescence, however none of the primer sets show any thing in the melting curve analysis.

qPCRs – BB & DH cDNA (from 20091223)

qPCR was set up on these cDNAs using the following primers:

AM861391.p.cg.6 (“BDEF”, “Big Defensin”) – This was upregulated in DH SOLiD data.

AM904566.p.cg.6 (“GNRR2″, “Gonadotropin-releasing hormone II receptor”) – This was upregulated in DH SOLiD data.

qPCR set up and plate layout can be found here.

Results:

qPCR Data File (Opticon): 20091228_102019.tad

Data workup: PROPS BB_DH Gene Expression Miner Workup

 

 

qPCR was set up on these cDNAs using the following primers:

CU988730.p.cg.6 (“TIMP3″, “Metalloprotease inhibitor 3″) – This was upregulated in DH SOLiD data.

CU990442.p.cg.6 (“CALL”, “Calmodulin-like protein”) – This was upregulated in DH SOLiD data.

qPCR set up and plate layout can be found here.

Results:

qPCR Data File (Opticon): 20091228_135507.tad

Data workup: PROPS BB_DH Gene Expression Miner Workup

 

 

qPCR was set up on these cDNAs using the following primers:

CU994646.p.cg.6 (“CATL”, “Cathepsin L”) – This was upregulated in DH SOLiD data.

ES789598.p.cg.6 (“GSTA”, “Glutathione S-transferase A”) – This was upregulated in DH SOLiD data.

qPCR set up and plate layout can be found here.

Results:

qPCR Data File (Opticon): 20091228_165801.tad

Data workup: PROPS BB_DH Gene Expression Miner Workup

qPCRs – BB & DH cDNA (from yesterday)

qPCR was set up on these cDNAs using the following primers:

EW778094 (“Ficolin 3″) –

AJ422120.p.cg.6 (“MDR49″, “Multidrug resistance protein homolog 49″) – This was upregulated in DH SOLiD data.

qPCR set up and plate layout can be found here.

Results:

qPCR Data File (Opticon): 20091224_092959.tad

Data workup: PROPS BB_DH Gene Expression Miner Workup

Preliminary results show that the EW778094 primer set looks bad; bad fluorescence profile and poor melt curves. Primers may require optimization.

 

 

qPCR was set up on these cDNAs using the following primers:

AM855874.p.cg.6 (“CP17A”, “Steroid 17-alpha-hydroxylase/17″) – This was upregulated in DH SOLiD data.

AM857078.p.cg.6 (“C1QT4″, Complement C1q tumor necrosis factor-related protein 4″) – This was upregulated in DH SOLiD data.

qPCR set up and plate layout can be found here.

Results:

qPCR Data File (Opticon): 20091224_125230.tad

Data workup: PROPS BB_DH Gene Expression Miner Workup

 

qPCR – Tim’s adult gigas challenge cDNA (from today)

Set up qPCR with EF1 primers and IL17 Internal primers. Plate layout/setup is here. Note: gDNA sample used as a “positive” control will NOT amplify with the EF1 primers.

Results: Processed with PCR Miner. Normalized to EF1. Standard Error bars. Here is spreadsheet with workup.

IL17 Internal:

No significant differences between any treatments.

qPCR – Tim’s adults gigas challenge re-DNased RNA (from today)

Performed qPCR using q18s primers on re-DNased RNA (1:100 dilution to match final concentration of template after making cDNA). qPCR set up and plate layout are here.

gDNA dilutions were used as positive controls. gDNA = BB11 (0.49ug/uL) from 20090519. Used 5uL of 1:10, 1:100 and 1:000 dilutions.

Results: re-DNase-ing the samples seems to have worked. Positive controls are the only samples to come up. Will proceed to making cDNA.