Total RNA was isolated using RNA-Bee reagent (Tel-Test) from FACS sorted 15.5 dpc male germ cells, followed by deoxyribonuclease I treatment. Libraries were prepared by the VAI Genomics Core from 500 ng of total RNA using the KAPA RNA HyperPrep Kit (Kapa Biosystems, Wilmington, MA, USA). Ribosomal RNA material was reduced using the QIAseq FastSelect –rRNA HMR Kit (QIAGEN, Germantown, MD, USA). RNA was sheared to 300 to 400 bp. Before PCR amplification, cDNA fragments were ligated to IDT for Illumina TruSeq UD indexed adapters (Illumina Inc., San Diego, CA) and amplified with 8 cycles of PCR. Quality and quantity of the finished libraries were assessed using a combination of Agilent DNA High Sensitivity chip (Agilent Technologies Inc.) and QuantiFluor dsDNA System (Promega Corp., Madison, WI, USA). Individually indexed libraries were pooled, and 50-bp, paired-end sequencing was performed on an Illumina NovaSeq 6000 sequencer to an average depth of 100 million raw paired reads per transcriptome. Base calling was done by Illumina RTA3, and output of noncrystallographic symmetry was demultiplexed and converted to FastQ format with Illumina Bcl2fastq v1.9.0.
Truseq ud indexed adapters
Manufactured by Illumina
Sourced in United States
Illumina TruSeq UD Indexed adapters are a set of oligonucleotide sequences designed to be ligated to DNA fragments during library preparation for sequencing on Illumina platforms. These adapters enable unique dual indexing of each sample, allowing for accurate sample identification and multiplexing of numerous samples in a single sequencing run.
Automatically generated - may contain errors
Lab products found in correlation
Agilent dna high sensitivity chip, by Agilent Technologies (7 mentions)
Quantifluor dsdna system, by Promega (7 mentions)
Novaseq 6000 sequencer, by Illumina (6 mentions)
Bcl2fastq v1, by Illumina (5 mentions)
Kapa rna hyperprep kit, by Roche (5 mentions)
Qiaseq fastselect rrna hmr kit, by Qiagen (5 mentions)
Rna bee reagent, by Tel-Test (2 mentions)
Kapa mrna hyperprep kit v4.17, by Roche (1 mentions)
Kapa rna hyperprep kit with riboseerase v1.16, by Roche (1 mentions)
Qiaseq fastselect 5s 16s 23s kit, by Qiagen (1 mentions)
Novaseq 6000, by Illumina (1 mentions)
Bcl2fastq, by Illumina (1 mentions)
7 protocols using truseq ud indexed adapters
1
Single-Cell RNA-Seq of 15.5 dpc Germ Cells
2
Single-Cell RNA-Seq of 15.5 dpc Germ Cells
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Total RNA was isolated using RNA-Bee reagent (Tel-Test) from FACS sorted 15.5 dpc male germ cells, followed by deoxyribonuclease I treatment. Libraries were prepared by the VAI Genomics Core from 500 ng of total RNA using the KAPA RNA HyperPrep Kit (Kapa Biosystems, Wilmington, MA, USA). Ribosomal RNA material was reduced using the QIAseq FastSelect –rRNA HMR Kit (QIAGEN, Germantown, MD, USA). RNA was sheared to 300 to 400 bp. Before PCR amplification, cDNA fragments were ligated to IDT for Illumina TruSeq UD indexed adapters (Illumina Inc., San Diego, CA) and amplified with 8 cycles of PCR. Quality and quantity of the finished libraries were assessed using a combination of Agilent DNA High Sensitivity chip (Agilent Technologies Inc.) and QuantiFluor dsDNA System (Promega Corp., Madison, WI, USA). Individually indexed libraries were pooled, and 50-bp, paired-end sequencing was performed on an Illumina NovaSeq 6000 sequencer to an average depth of 100 million raw paired reads per transcriptome. Base calling was done by Illumina RTA3, and output of noncrystallographic symmetry was demultiplexed and converted to FastQ format with Illumina Bcl2fastq v1.9.0.
3
RNA-seq Library Preparation and Sequencing
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Libraries were prepared by the Van Andel Institute Genomics Core from 500 ng of total RNA using the KAPA mRNA Hyperprep kit (v4.17) (Kapa Biosystems, Wilmington, MA United States). RNA was sheared to 300–400 bp. Prior to PCR amplification, cDNA fragments were ligated to IDT for Illumina TruSeq UD Indexed adapters (Illumina Inc., San Diego CA, United States). Quality and quantity of the finished libraries were assessed using a combination of Agilent DNA High Sensitivity chip (Agilent Technologies, Inc.), QuantiFluor® dsDNA System (Promega Corp., Madison, WI, United States), and Kapa Illumina Library Quantification qPCR assays (Kapa Biosystems). Individually indexed libraries were pooled and 50 bp, paired-end sequencing was performed on an Illumina NovaSeq6000 sequencer using an S2 sequencing kit (Illumina Inc., San Diego, CA, United States) to an average depth of 45M reads per sample. Base calling was done by Illumina RTA3 and output of NCS was demultiplexed and converted to FastQ format with Illumina Bcl2fastq v1.9.0.
4
RNA Sequencing Library Preparation and Sequencing
5
RNA-seq Library Preparation and Sequencing
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Libraries were prepared by the Van Andel Genomics Core from 500 ng of total RNA using the KAPA RNA HyperPrep Kit (Kapa Biosystems, Wilmington, MA USA). Ribosomal RNA material was reduced using the QIAseq FastSelect –rRNA HMR Kit to remove cytoplasmic and mitochondrial rRNAs and QIAseq FastSelect –5S/16S/23S Kit to remove pan bacterial rRNAs (Qiagen, Germantown, MD, USA). RNA was sheared to 300-400 bp. Prior to PCR amplification, cDNA fragments were ligated to adapters for Illumina TruSeq UD Indexed adapters (Illumina Inc, San Diego CA, USA). Quality and quantity of the finished libraries were assessed using a combination of Agilent DNA High Sensitivity chip (Agilent Technologies, Inc.), QuantiFluor® dsDNA System (Promega Corp., Madison, WI, USA), and Kapa Illumina Library Quantification qPCR assays (Kapa Biosystems). Barcodes that are unique to each sample (from each patient) were added to each DNA fragment; and resulting DNA libraries were pooled (combined) in the sequencing run. The sequencing was performed on an Illumina NovaSeq6000 targeting depth of 100 M per sample with 100 bp, paired-end configuration. Base calling was done by Illumina Real Time Analysis and output of NextSeq Control Software was demultiplexed and converted to Fastq format with Illumina Bcl2fastq v1.9.0.
6
RNA-seq Library Preparation and Sequencing
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Libraries were prepared by the Van Andel Institute Genomics Core (RRID:SCR_022913) from 500 ng of total RNA using the KAPA RNA HyperPrep Kit (Kapa Biosystems, Wilmington, MA, USA). Ribosomal RNA material was reduced using the QIAseq FastSelect–rRNA HMR Kit (QIAGEN, Germantown, MD, USA). RNA was sheared to 300 to 400 bp. Before PCR amplification, cDNA fragments were ligated to IDT for Illumina TruSeq UD Indexed adapters (Illumina Inc., San Diego CA, USA). Quality and quantity of the finished libraries were assessed using a combination of the Agilent DNA High Sensitivity chip (Agilent Technologies Inc.) and QuantiFluor dsDNA System (Promega Corp., Madison, WI, USA). Individually indexed libraries were pooled and 50 bp, paired-end sequencing was performed on an Illumina NovaSeq6000 sequencer to an average depth of 50 M raw paired-reads per transcriptome. Base calling was done by Illumina RTA3, and output of NCS was demultiplexed and converted to FastQ format with Illumina Bcl2fastq (v1.9.0).
7
RNA-seq Library Preparation and Sequencing
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+ Expand
Libraries were prepared by the Van Andel Genomics Core from 500 ng of total RNA using the KAPA RNA HyperPrep Kit (Kapa Biosystems, Wilmington, MA USA). Ribosomal RNA material was reduced using the QIAseq FastSelect -rRNA HMR Kit (Qiagen, Germantown, MD, USA). RNA was sheared to 300-400 bp. Before PCR amplification, cDNA fragments were ligated to IDT for Illumina TruSeq UD Indexed adapters (Illumina Inc, San Diego CA, USA). Quality and quantity of the finished libraries were assessed using a combination of Agilent DNA High Sensitivity chip (Agilent Technologies, Inc.), QuantiFluor ® dsDNA System (Promega Corp., Madison, WI, USA), and Kapa Illumina Library Quantification qPCR assays (Kapa Biosystems). Individually indexed libraries were pooled and 50 bp, paired-end sequencing was performed on an Illumina NovaSeq6000 sequencer to an average depth of 30M raw paired reads per transcriptome. Base calling was done by Illumina RTA3 and the output of NCS was demultiplexed and converted to FastQ format with Illumina Bcl2fastq v1.9.0.
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