Apr 18, 2024

Public workspace10x Protocols: Chromium Next GEM Single Cell 5' -- University of Minnesota TMCsTMCs (CG000331 Rev E)

DOI
dx.doi.org/10.17504/protocols.io.5qpvokn89l4o/v1
  • Laura J Niedernhofer1,
  • David A Bernlohr1
  • 1University of Minnesota, Minneapolis, MN
Allie Pybas
Open access
DOI: dx.doi.org/10.17504/protocols.io.5qpvokn89l4o/v1
Protocol Citation: Laura J Niedernhofer, David A Bernlohr 2024. 10x Protocols: Chromium Next GEM Single Cell 5' -- University of Minnesota TMCsTMCs (CG000331 Rev E). protocols.io https://dx.doi.org/10.17504/protocols.io.5qpvokn89l4o/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: April 02, 2024
Last Modified: April 18, 2024
Protocol Integer ID: 97664
Funders Acknowledgement:
NIH
Grant ID: 5U54AG076041-03
NIH
Grant ID: 5U54AG079754-02
Abstract
 Following single-cell dissociation or nuclei isolation, the Next GEM 5' assay uses microfluidics to partition and assign cell- or nuclei-specific barcodes to transcript cDNA at the 5' end. Barcoded cDNA is prepared with adaptors for sequencing by synthesis (SBS). The following protocol has been used at the University of Minnesota TMCs in collaboration with the University of Minnesota Genomics Center. The following protocol has been adapted from protocols developed by 10x Genomics and Illumina to be used at the University of Minnesota TMCs in collaboration with the University of Minnesota Genomics Center. These protocols are owned by their respective companies and are subject to periodic revision.
Tissue Preparation
Tissue Preparation
Complete single cell or nuclei isolation prior to starting this protocol
Library Preparation
Library Preparation
Download CG000331_Rev-E.pdfCG000331_Rev-E.pdf4.3MB

Note
Sequence with the read format 29,10,10,89


Note
Sequencers used at UMN Genomics Center:
  • Illumina NextSeq 2000
  • Illumina NovaSeq 6000
  • Illumina NovaSeq X Plus

FASTQ Generation
FASTQ Generation
BCL data from Illumina sequencer is demultiplexed and converted into FASTQ format using bcl2fastq version 2.20.0