User:Karmella Haynes/Notebook/Polycomb project/2010/06/15

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 * style="background-color: #F2F2F2" align="center"|  |Main project page


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06/15/10

 * &#x2713; Western: protein prep 126-1, 126-3, 132-8 -/+ dox, 4 days (from 6-well)
 * &#x2713; RT-PCR (set #1): 126-1, 126-3, 132-8, 154-2, FTRx -/+ dox, 4 days RNA preps (from 6-well); cDNA
 * &#x2713; Cell culture: split cells 1:10 (10 cm plates)
 * &#x2713; RT-PCR (set #2): start new 6-well cultures (-/+ dox) for 126-1, 126-3, 132-8, 154-2, FTRx

RNA Preps/ cDNA synth.

> RNA Preps --> Use 1 mL TriZol/ well (6-well dishes) --> Resuspend RNA pellet in 10 μL THE RNA Storage Solution

> oligo(dT) Primer annealing

> cDNA synthesis mix --> 10 reactions total

--> Add 10 μL mix to each annealing rxn. --> 50°C/ 50 min., 80°C/ 5 min., ice --> Add 1.0 μL RNase H, 37°C/ 20 min. --> Store at -20°C


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