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Scun22 PCRs from 7 and 10 February
- These are from the PCRs that Rulon and I demo'd to the students
- In both cases, used Scun22 primers, run on both Scoc and Scun DNA.
- We used the same PCR sheet, but Rulon put his samples in A12-H12 of his PCR plate. Same reaction volumes used.
- Rulon PCR'd 7 Feb 2011; Tara PCR'd 10 Feb 2011
Scun22 Agarose Gel
- Pour small Gel: 50 mL 1x TAE + 1 g agarose + 5 µL GelRed
- Load: 5µL Orange/Blue loading dye + 2 µL PCR product
- This is the first time I'm trying the Orange/Blue Loading Dye (Orange G and Xylene Cyanol FF), stolen from Joe Newsome's excess stock
- Run: 80V, 40 min
- ----
 - oops, can't see product because I used the 6x loading dye like an idiot
- did dye:DNA 1:5 instead of DNA:dye 1:5
Scun22 Agarose Gel Corrected
- Pour small Gel: 50 mL 1x TAE + 1 g agarose + 5 µL GelRed
- Load: 1µL Orange/Blue loading dye + 5 µL PCR product
- Using the Orange/Blue Loading Dye (Orange G + Xylene Cyanol FF + Bromophenol Blue), stolen from Joe Newsome's excess stock
- Run: 90V, 20 min
- -
 - ---
 - Only two samples amplified:
- Tara's Scoc (4th replicate) - primary band at expected ~450 bp, but with mispriming
- Tara's Scun (1st replicate) - primary band at expected ~450 bp
- This pattern (Scoc dirty, Scun clean) is consistent with results from 14 January
Next Steps
- Optimize by varying MgCl2, convert to standard thermal cycling profile ... full gradient PCR (temp and Buffer)
- Optimize by converting to standard thermal cycling profile
- Try to get their temperatures to match!
Meeting with Rulon
- completed online CSU-approved defensive driving course - send certificate of completion to DPS
 - completed two other forms ... keep with department? Ask Medora
- CA DFG SCP - send email to Randi Logsdon requesting status update; no response, yet
- pipet and tips
- prefer to order GeneMate, even though it would require stocking different tips
- Rulon gives go-ahead
Field Work
- for use in the field (to ensure each sample has a unique ID)
- MMDDYY initial A, B, C ...
- TP3-1-10
- TP3-11-15
- TP1-1-10
- in degrees and minutes (seconds as decimals of minute)
- split into 2: one for DNA, one for isotope (no alcohol)
- lizard = toe and tail
- snake = scale
- print color copies of maps, in plastic sleeve
- county, site, array
- get copies of Rulon's permits
- email Carlton for full database (so we can share our stuff with USGS at end)
- get copies from Rulon
- data sheet, toe clip, scale clip
- Pesola Scales (10g, 20g, 60g, 300g)
- Rulers (stick and tape)
- lab pens (2)
- bunch o' baggies
- forceps
- scissors
- QuikStop
- alcohol wipes
- ethanol tubes
- hand Wipies
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Tara's Lab Notebook
