User:Karmella Haynes/Notebook/Polycomb project/2010/12/07

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12/07/10

 * &#x2713; ChIP qPCR: continue with MMP12 primers
 * &#x2713; Cell culture: expand KAH126-1, 128-8.3, 129 to 75 cm flasks (130-4 did not grow)

ChIP qPCR > Set up each reaction in triplicate > Templates (use 2.0 μL): > Primers (21 rxns pre primer pair): --> Plate 2 --> 750 nM primer mix = 3 μL 100 μM each primer + 394 μL H2O
 * FTRx dx input, pos
 * FTRx dx H3K27me3 IP, uk
 * FTRx dx myc, neg
 * FTRx fx input, pos
 * FTRx fx H3K27me3 IP, uk
 * FTRx fx myc, neg
 * 0 template
 * 1) MMP12 A3
 * 2) MMP12 B2
 * 3) MMP12 C2
 * 4) MMP12 D3

--> Aliquot 39.0 primer mix into 1st well of each triplicate --> Add 6.0 (2.0 x 3) DNA to 39.0 primer mix --> Aliquot 15.0 rxn mix to other 2 wells in each 3x set

Bio-Rad CFX96 qPCR (Kirschner lab) --> Use Bio-Rad 96-well low profile plate MLL-9601 + Microseal "B" film
 * 95°C/ 5 min.
 * [95°C/ 15 sec, 57°C/ 15 sec, 72°C/ 15 sec] x45
 * Melt curve range 57°C -> 95°C/ 0.5°C per step


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