Kapa library quantification dna standards 1 5

Libraries from all experiments were profiled, undiluted, on an Agilent TapeStation 4200 with the High Sensitivity D1000 Reagents (#5067–5585). Libraries to be sequenced were diluted to 1/10,000 concentration by serial 1/100 dilutions and quantified by qPCR with the primers 5′-AATGATACGGCGACCACCGA-3′ and 5′-CAAGCAGAAGACGGCATACGA-3′ at 450nM each, using PowerUp SYBR Green Master Mix (Thermo Fisher Scientific #A25742) and a program of 2min at 50 °C, 2min at 95 °C, then 40 cycles of 15s at 95 °C, 1min at 60 °C. The KAPA Library Quantification DNA Standards #1–5 (Roche #07960387001) were used for the standard curve. The qPCR measurements were scaled by the qPCR’s dilution factor and the TapeStation’s average library molecule lengths to calculate the library molarities used for pooling.

Libraries from all experiments were profiled, undiluted, on an Agilent TapeStation 4200 with the High Sensitivity D1000 Reagents (#5067-5585). Libraries to be sequenced were diluted to 1/10,000 concentration by serial 1/100 dilutions and quantified by qPCR with the primers 5′-AATGATACGGCGACCACCGA-3′ and 5′-CAAGCAGAAGACGGCATACGA-3′ at 450 nM each, using PowerUp SYBR Green Master Mix (#A25742; Thermo Fisher Scientific) and a program of 2 min at 50°C, 2 min at 95°C, then 40 cycles of 15 s at 95°C, 1 min at 60°C. The KAPA Library Quantification DNA Standards #1–5 (#07960387001; Roche) were used for the standard curve. The qPCR measurements were scaled by the qPCR’s dilution factor and the TapeStation’s average library molecule lengths to calculate the library molarities used for pooling.