Beckman Coulter
SPRIselect
B23318
SPRIselect reagent was used for selecting 550 to 600 bp adapter ligated whole genome sequencing libraries after PCR amplification with i5 and i7 illumina indexing primers. Post size selection libraries were analyzed by Agilent 2100 High sensitivity DNA kit and indeed the libraries were 560 bp in size.
Size selection and PCR clean up of whole genome sequencing libraries
Genomic DNA sheared, adapter ligated and PCR amplified using Illumina indexing primers
Genomic DNA which are adapter ligated and amplified by PCR using Illumina i7 and i5 indexing primers were size selected for 550-600 bp size fragments (insert plus adapter ligated) using SPRIselect reagent. First selection was at 0.2X concentration of SPRIselect to PCR amplified libraries. Second size selection at 0.1X concentration of SPRIselect reagent which actually contains desired fragment libraries. After second selection, DNA libraries were washed by 2 washes with 80% ethanol while SPRIselect beads containing libraries still on magnetic rack. Finally eluted with 33 microliter of nuclease free water.
Mixing of DNA libraries with SPRIselect reagent should be done thoroughly without introducing much air baubles. SPRIselect reagent before using should be allowed to normalise at room temperature for 30 minutes.
Post size selection libraries analysed by Agilent 2100 high sensitivity DNA kit revealed size of the DNA libraries in the range of 550 to 580 bp.
N/A
Simple and easy to follow protocol.
None
Must usable and recommended reagent for preparation of whole genome sequencing libraries.