Duplicates of earlier qPCRs.
Primers: Cg_P450 and TNFRAF_5’/3′.
qPCR set up and plate layout are here.
Results:
Duplicates of earlier qPCRs.
Primers: Cg_IkB_F997, R1213 and Cg_Prx6_F270, R439.
qPCR set up and plate layout can be found here.
Results:
SOLiD ePCR – Herring fragmented cDNA library: 3LHSITK09 (from 20091209)
Processed herring fragmented cDNA library 3LHSITK09 (88.ng/uL) according to the ABI “Templated Bead Preparation Guide” following the “full-scale” protocol. Made a 1:1000 dilution (1uL library, 999uL 1x Low TE) = 88.5pg/uL. Mixed 67.8uL of this diluted sample with 32.2uL 1x Low TE to get a final concentration of 60pg/uL (500pM, according to ABI protocol). Oil phase used was previously prepared by Jesse (Seeb lab) in mid-December 2009. This oil phase is stable for 2 months @ 4C.
ePCR was started (Rhonda will put plate in fridge for storage O/N) and the rest of the procedure will be finished tomorrow.
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
Alaska sockeye salmon sampling (with Seebs): Family #13
Juvenile sockeye salmon were subjected to initial heat stress of 18C. 10 fish were weighed, measured and then snap frozen in LN2. Samples were transferred to a freezer box labelled “AL Sockey Family #13″ and stored @ -80C. Here is the spreadsheet with all the pertinent info.
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