Caenorhabditis elegans genomic sequence, jf54 mutant (rmh-1) cross sample 30405
50ul of frozen worm pellet was lysed in 450μl lysis buffer with 30μl of proteinase K (20mg/μl) for 1hour at 60°C. Proteinase K was inactivated at 95°C for 15min. RNAs were eliminated by adding 5ul of RNAse A (10mg/μl) to the sample and incubated at 42°C for 2 hours. 80μl of 5M NaCl and 80μl of CTAB/Nacl were added and samples were incubated at 65°C for 10min. After phenol/chloroform extraction, DNA was precipitated with ethanol and re-suspended in water. DNA sample were then sheared using a S220 ultrasonicator (Covaris). Measure concentration of DNA was done on Fluorospectrometer, 5ng of DNA in 40µl was needed for sample preparation which follows the next steps: End Repair, Ligate Adapters to DNA Fragments, Size select the library, Purification using QIAquick Gel Extraction Kit, Enrich the adapter-modified DNA Fragments by 18 PCR cycles, Library Validation using the Agilent Bioanalyzer and Check of concentration using qPCR. Samples were then sequenced on an Illumina HiSeq2500 with 125 PE-sequencing.
MFPL
MUT_jf54_Celegans_N2xHw_cross_30405