AB SOLiD 4 System sequencing; Identification of Elg1 interaction partners and effects on post-replication chromatin re-formation
Identification of Elg1 interaction partners and effects on post-replication chromatin re-formation
All strains were grown on YPD plates at 30 degrees Celsius prior to inoculation for ChIP-Seq experiment. OD600 0.25 cells were arrested in S-phase by addition of alpha factor. After 2 hours, cells were released into S-phase by adding Pronase at 23 degrees Celsius into YPD medium containing 0.2M Hydroxyurea and incubated for 60 minutes. Formaldehyde was then added to a final concentration of 1%. Cells were then incubated at room temperature for 20 minutes and then in cold room overnight. Cells were then washed with ice cold 1X PBS thrice. Cell pellet was frozen at -80 degrees Celsius until further processing. Cells were disrupted using Multi-beads shocker (MB400U, Yasui Kikai) Anti-HA monoclonal antibody HA.11 (16B12) was used for Immunoprecipitation.  DNA from Input and ChIP fractions were sheared to an average size of 150 bp using a Covaris Ultra. DNA was then subjected to end repair and ligation of primers before eight rounds of amplification and the samples were sequenced using ABI Solid version 4.0 system according to manufacturer instructions.
Experimental Factor: genotype
wild type genotype - Mata arg4::natMX4 his31 leu20 lys20 ura30 RTT106-6HA::hphNT1
Experimental Factor: stimulus
-factor block
Experimental Factor: time
G1 control
Experimental Factor: protocol
ChIP Seq
Experimental Factor: immunoprecipitate
input DNA