Total RNA from gastrocnemius muscle was isolated using the RNeasy Mini Kit (74,106, Qiagen). Quantification of RNA was determined using a spectrophotometer (Nanodrop 8000, Thermo Scientific), and integrity was measured using a Bioanalyzer 2100 (Agilent). Total RNA sequencing libraries were prepared by the Single-Cell Omics platform at the Center for Basic Metabolic Research using the Illumina TruSeq Stranded total RNA Gold protocol (Illumina). The total RNA (380 ng) was depleted of rRNA by RiboZero beads, fragmented, and cDNA was synthesized using SuperScript III Reverse Transcriptase (Thermo Fisher Scientific). cDNA was adenylated to prime for adapter ligation, and after a clean-up using AMPure beads (Beckman coulter), DNA fragments were amplified using PCR, followed by a final clean-up. Libraries were quality controlled using a Bioanalyzer instrument (Agilent Technologies) and subjected to 38-bp paired-end sequencing on a NextSeq 500 system (Illumina).
Truseq stranded total rna gold protocol
Manufactured by Illumina
Sourced in United States
The TruSeq Stranded total RNA Gold protocol is a library preparation kit designed for RNA sequencing. It enables the conversion of total RNA into a library of template molecules suitable for subsequent cluster generation and sequencing. The protocol is optimized for stranded RNA-Seq, which preserves the strand orientation of the original RNA molecules.
Automatically generated - may contain errors
Lab products found in correlation
Turbo dna free kit, by Thermo Fisher Scientific (4 mentions)
Ribozero beads, by Thermo Fisher Scientific (2 mentions)
Superscript 3 reverse transcriptase, by Thermo Fisher Scientific (2 mentions)
Rneasy mini kit, by Qiagen (2 mentions)
Bioanalyzer, by Agilent Technologies (2 mentions)
Ampure bead, by Beckman Coulter (2 mentions)
2100 bioanalyzer, by Agilent Technologies (2 mentions)
Trizol, by Thermo Fisher Scientific (2 mentions)
Nanodropnd 6000, by Thermo Fisher Scientific (2 mentions)
Allprep dna rna mini kit, by Qiagen (2 mentions)
Nextseq 500 system, by Illumina (1 mentions)
Nanodrop 8000, by Thermo Fisher Scientific (1 mentions)
Humanht 12 v4, by Illumina (1 mentions)
Human ht 12 v4.0 beadchips, by Illumina (1 mentions)
Hiseq 4000, by Illumina (1 mentions)
Hiseq 2000, by Illumina (1 mentions)
Agilent 2100 bioanalyzer, by Agilent Technologies (1 mentions)
4 protocols using truseq stranded total rna gold protocol
1
Gastrocnemius Muscle RNA Sequencing
2
Total RNA-Seq Library Preparation
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Total RNA was isolated using Trizol (Thermo Fisher Scientific) followed by purification with an RNeasy mini kit (Qiagen) according to the manufacturer's protocol. Total RNA-Seq libraries were prepared using the Illumina TruSeq Stranded total RNA Gold protocol (Illumina). The total RNA (500 ng) was depleted of rRNA by RiboZero beads, fragmented, and cDNA was synthesized using SuperScript III Reverse Transcriptase (Thermo Fisher Scientific). cDNA was adenylated to prime for adapter ligation, and after a cleanup using AMPure beads (Beckman Coulter), DNA fragments were amplified using PCR followed by a final cleanup. Libraries were quality controlled using a Bioanalyzer instrument (Agilent Technologies).
3
Endometrial Transcriptomic Profiling Protocol
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Of the 358 endometrial samples, 290 had Illumina Human HT-12 v4.0 performed and 266 underwent RNA-seq (198 samples had both techniques performed). Total RNA was isolated from endometrial samples using the Allprep DNA/RNA Mini Kit (Qiagen, CA) as per the manufacturer’s instructions29 (link). Briefly, RNA quality was checked using a Bioanalyzer 2100 (Agilent Technologies, CA) and RNA concentration was measured using a NanoDropND-6000 (Thermo Fisher Scientific, USA). All samples were high quality with an RNA integrity number greater than 8. Expression profiles in endometrial tissue were generated by hybridizing 750 ng of cRNA to Illumina Human HT-12 v4.0 Beadchips.
RNA samples were treated with Turbo DNA-free kit (Thermo Fisher Scientific, USA) prior to RNA-seq library generation19 (link). Stranded RNA-seq libraries were prepared using the Illumina TruSeq Stranded Total RNA Gold protocol which includes ribosomal depletion (Illumina, USA).
RNA samples were treated with Turbo DNA-free kit (Thermo Fisher Scientific, USA) prior to RNA-seq library generation19 (link). Stranded RNA-seq libraries were prepared using the Illumina TruSeq Stranded Total RNA Gold protocol which includes ribosomal depletion (Illumina, USA).
4
Endometrial Transcriptome Profiling by RNA-Seq
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Total RNA was isolated from endometrial biopsies stored in RNAlater using the Allprep DNA/RNA Mini Kit (Qiagen, USA) as per the manufacturer’s instructions and isolated RNA samples were treated with the Turbo DNA-free kit (Thermo Fischer Scientific, USA). RNA quality was confirmed using the Agilent Bioanalyzer 2100 (Agilent Technologies, USA) with RNA integrity number (RIN) cut off values set at above 8 for inclusion and final RNA concentrations determined by the Nanodrop ND -6000 (Thermo Fisher Scientific, USA). Stranded RNA-sequencing (RNA-seq) libraries were prepared with the Illumina TruSeq Stranded Total RNA Gold protocol incorporating ribosomal depletion (Illumina, USA). The resulting libraries were pooled and sequenced with 75 bp paired-end reads on the Illumina HiSeq 4000 to a mean depth of 37,490,673 for 178 samples and with 120 bp paired-end reads on an Illumina Hi Seq 2000 (Illumina, USA) for a mean depth of 40,818,062 reads for 28 samples.
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