User:Karmella Haynes/Notebook/PcTF Genomics/2011/12/13

12/13/11

• ✓ Illumina ChIp-seq prep: Pc-TF ChIP samples 1-6 (Day one)

Illumina ChIP-seq prep
> Follow manufacturer's protocol, with some modifications
> Samples (dated 12/20/10):

1. "32, 126-1 myc" (hPc-TF)
2. "33, 126-1 IgG" (hPc-TF mock)
3. "36, 132-8 myc" (TF)
4. "37, 132-8 myc" (TF mock)
5. "38, FTRx K27m" (plain cells)
6. "39, FTRx K27m" (plain cells mock)

Perform End Repair
Note: forgot to dilute Klenow 1:5 with water before use

--> Aliquot 20 μL master mix, add 30 μL ChIP DNA
--> 20°C/ 30 min. (Xiao's thermal cycler)
--> QIAquick PCR Purification, elute with 34 μL EB

Add 'A' Bases to the 3' End of the DNA Fragments

--> Aliquot 16 μL master mix into DNA
--> 37°C/ 30 min. (heat block)
--> Zymo clean and concentrator, elute with 10 μL st.dH₂O

Ligate Adapters to DNA Fragments
Note: Did not dilute the adapter oligo mix

--> Aliquot 20 μL to DNA samples
--> 15 min./ r.t.
--> Zymo clean and concentrator, elute with 10 μL st.dH₂O
--> Store at 4°C o/n