PacBio HiFi data were generated from the HG00733 lymphoblastoid cell line as previously described (G. A. Logsdon et al., 2021 (link)) with modifications. Briefly, DNA was extracted from 4.3×10^6 cells using the Monarch HMW DNA Extraction Kit for Cells and Blood (New England Biolabs) with 1400 rpm lysis speed. After UV absorption and fluorometric quantification (Qubit High Sensitivity DNA kit, Thermo Fisher) on the DS-11 FX instrument (Denovix) and evaluation of DNA integrity on FEMTO Pulse (Agilent), 12 μg of DNA was prepared for sequencing using Megaruptor 3 shearing (Diagenode, settings 19/31) and the Express Template Prep Kit v2 and SMRTbell Cleanup Kit v2 (PacBio). The library was size-selected on a PippinHT instrument (Sage Science) using a 15 kbp high-pass cut. Five SMRT Cell 8Ms were run on a Sequel II instrument using Sequel II chemistry C2.0/P2.2 with 30-hour movie times, 2-hour pre-extension, and adaptive loading targets of 0.8–0.85 (PacBio). Circular consensus calling was performed with CCS version 6.0.0 (SMRT Link v.10.1) and reads with estimated quality scores ≥Q20 were selected for downstream analysis.
Sequel 2 chemistry c2 0 p2
Manufactured by Pacific Biosciences
The Sequel II chemistry C2.0/P2.2 is a core component of the Sequel II System, a long-read sequencing platform developed by Pacific Biosciences. It is responsible for the key chemical reactions and processes involved in generating high-quality sequencing data.
Automatically generated - may contain errors
Lab products found in correlation
Ds 11 fx instrument, by DeNovix (2 mentions)
Monarch hmw dna extraction kit for cells and blood, by New England Biolabs (2 mentions)
Megaruptor 3 shearing, by Diagenode (2 mentions)
Express template prep kit v2 and smrtbell cleanup kit v2, by Pacific Biosciences (2 mentions)
Qubit high sensitivity dna kit, by Thermo Fisher Scientific (2 mentions)
Pippinht instrument, by Sage Science (2 mentions)
Femto pulse, by Agilent Technologies (2 mentions)
2 protocols using sequel 2 chemistry c2 0 p2
1
PacBio HiFi Sequencing of HG00733
2
HiFi Sequencing of HG00733 Cell Line
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PacBio HiFi data were generated from the HG00733 lymphoblastoid cell line as previously described (Logsdon et al. 2021 (link)) with modifications. Briefly, DNA was extracted from 4.3 × 106 cells using the Monarch HMW DNA Extraction Kit for Cells and Blood (New England Biolabs) with 1400 rpm lysis speed. After UV absorption and fluorometric quantification (Qubit High Sensitivity DNA kit, Thermo Fisher Scientific) on the DS-11 FX instrument (Denovix) and evaluation of DNA integrity on FEMTO Pulse (Agilent), 12 μg of DNA was prepared for sequencing using Megaruptor 3 shearing (Diagenode, settings 19/31) and the Express Template Prep Kit v2 and SMRTbell Cleanup Kit v2 (PacBio). The library was size-selected on a PippinHT instrument (Sage Science) using a 15 kbp high-pass cut. Five SMRT Cell 8Ms were run on a Sequel II instrument using Sequel II chemistry C2.0/P2.2 with 30-h movie times, 2-h pre-extension, and adaptive loading targets of 0.8–0.85 (PacBio). Circular consensus calling was performed with CCS version 6.0.0 (SMRT Link v.10.1) and reads with estimated quality scores ≥Q20 were selected for downstream analysis.
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