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=== Things to do upon receive the probe pool === '''1. Recommended by instruction in CHAPTER2:''' :Upon receipt, undertake the following steps to ensure the highest performance of the SeqCap Epi probe pool to avoid multiple freeze/thaw cycles or potential accidental contamination: # If frozen, thaw the tube of SeqCap Epi probe pool on ice. # Vortex the SeqCap Epi probe pool for 3 seconds. # Centrifuge the tube of SeqCap Epi probe pool at 10,000 x g for 30 seconds to ensure that the liquid is at the bottom of the tube before opening the tube. # Aliquot the SeqCap Epi probe pool into single-use aliquots (4.5 Β΅l/aliquot) in 0.2 ml PCR tubes (or 96-well plates if following the higher throughput protocol described in Appendix C) and store at -15 to -25Β°C until use. The presence of some residual volume after dispensing all single-use aliquots is normal. # When ready to perform the experiment, thaw the required number of single-use SeqCap Epi probe pool aliquots on ice. '''NOTE:''' The SeqCap Epi probe pool should not undergo multiple freeze/thaw cycles. To help ensure the highest performance of the SeqCap Epi probe pool, Roche NimbleGen recommends aliquoting the SeqCap Epi probe pool into single-use volumes to prevent damage from successive freeze/thaw cycles.<br> '''Label PCR tube:''' Hg19 UMR v1 141029 '''Location:''' -20C, in original box within Noi Library prep kit box. I got total '''16 tubes of 4.5ul aliquot'''. The instruction said it's normal to get extra volume of the probe. '''2. For sequencing library preparation to decode the sequences of the probe pool''' # Take a small aliquot of the left over probe pool after a single-use aliquots of 16 rxn for library prep using ClonTech DNA SMRT ChIP-Seq kit <br> * There is ~2ul of the probe left in the tube after aliquot for single-use. I added 3ul of H2O to increase the volume to 5ul. I then took 1ul for Qubit ssDNA quantification. * Qubit result = 5.85ng/ul --> original concentration from Nimblegen should be ~ 14.62ng/ul. * I use 10ng or 1.71ul of a diluted probe for DNA SMART ChIP seq experiment. This amount is the maximum amount recommended by the kit (between 100pg-10ng of dsDNA or ssDNA) ---- ----
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