Approximately 10 g and 2 g of sediment were used to extract DNA and RNA using the DNeasy PowerMax Soil kit (QIAGEN) and RNeasy PowerSoil kit (QIAGEN) kits, respectively. DNase treatment was conducted on extracted RNA by using the TURBO DNA-free kit (Invitrogen). This was followed by ribosomal RNA depletion with the RiboMinus Transcriptome Isolation Kit (ThermoFisher Scientific). A 2100 Bioanalyzer (Agilent) was used to confirm that the RNA samples were free of DNA contamination. Library preparation of DNA and RNA samples were prepared with the ThruPLEX DNA-seq (Rubicon Genomics) and TruSeq RNA Library Prep v2 (without the poly-A selection step, Illumina) kits, respectively. The Illumina NovaSeq6000 platform was used to sequence DNA and RNA, with one S2 and S4 lane used for the DNA and RNA samples (a paired-end 2 × 150 bp setup), respectively. All samples were sequenced at the Science for Life Laboratory, Stockholm. The sequences are available in the NCBI BioProject repositories, PRJNA541421 (DNA) and PRJNA54 1422 (RNA).
Truseq rna library prep v2
Manufactured by Illumina
The TruSeq RNA Library Prep v2 is a sample preparation kit designed for constructing RNA sequencing libraries. It enables the generation of high-quality cDNA libraries from total RNA samples.
Automatically generated - may contain errors
Lab products found in correlation
Ribominus transcriptome isolation kit, by Thermo Fisher Scientific (3 mentions)
Turbo dna free kit, by Thermo Fisher Scientific (3 mentions)
Thruplex dna seq, by Takara Bio (3 mentions)
2100 bioanalyzer, by Agilent Technologies (2 mentions)
Rneasy powersoil kit, by Qiagen (2 mentions)
Dneasy powermax soil kit, by Qiagen (2 mentions)
Novaseq 6000 platform, by Illumina (1 mentions)
Nanodrop one, by Thermo Fisher Scientific (1 mentions)
Hiseq 4000, by Illumina (1 mentions)
Kapa hyper prep, by Roche (1 mentions)
Hiseq 2000 sequencer, by Illumina (1 mentions)
Direct zol kit, by Zymo Research (1 mentions)
Novaseq platform, by Illumina (1 mentions)
Novaseq 6000 s2, by Illumina (1 mentions)
Novaseq 6000 s4, by Illumina (1 mentions)
5 protocols using truseq rna library prep v2
1
Metagenomic DNA and RNA Extraction
2
DNA and RNA Extraction and Sequencing
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
DNA and RNA were extracted from one and two filters, respectively,
with the FastDNA SPIN Kit for the soil filter and two with the QIAGEN
RNeasy mini kit. Blanks were used during DNA extractions (exB1, exB2).
The TURBO DNA-free kit (Invitrogen) was used to DNase treat extracted
RNA and remove DNA contamination and afterward controlled for residual
DNA by a 30-cycle polymerase chain reaction (PCR) with 16S rDNA primers
(27F and 1492R) including Milli-Q as negative and DNA from Escherichia coli as positive controls. This was followed
by bacterial ribosomal RNA depletion using a RiboMinus Transcriptome
Isolation Kit (Thermo Fisher Scientific). The final quantity and quality
of extracted nucleic acids were measured on a NanoDrop One spectrophotometer
(Thermo Fisher Scientific) and a qubit. Library preparation of DNA
and RNA for sequencing was prepared with the ThruPLEX DNA-seq (Rubicon
Genomics) and TruSeq RNA Library Prep v2 (Illumina) kits, respectively.
The DNA and RNA samples were each sequenced with a paired-end 2 ×
150 bp setup at the Science for Life Laboratory (Stockholm) at the
SNP&SEQ Technology Platform over an entire Illumina NovaSeq6000
SP4 flow cell. The raw data are accessible at NCBI (SRA accession:
PRJNA627699). The bioinformatic procedure is described in detail in
theSupporting Information .
with the FastDNA SPIN Kit for the soil filter and two with the QIAGEN
RNeasy mini kit. Blanks were used during DNA extractions (exB1, exB2).
The TURBO DNA-free kit (Invitrogen) was used to DNase treat extracted
RNA and remove DNA contamination and afterward controlled for residual
DNA by a 30-cycle polymerase chain reaction (PCR) with 16S rDNA primers
(27F and 1492R) including Milli-Q as negative and DNA from Escherichia coli as positive controls. This was followed
by bacterial ribosomal RNA depletion using a RiboMinus Transcriptome
Isolation Kit (Thermo Fisher Scientific). The final quantity and quality
of extracted nucleic acids were measured on a NanoDrop One spectrophotometer
(Thermo Fisher Scientific) and a qubit. Library preparation of DNA
and RNA for sequencing was prepared with the ThruPLEX DNA-seq (Rubicon
Genomics) and TruSeq RNA Library Prep v2 (Illumina) kits, respectively.
The DNA and RNA samples were each sequenced with a paired-end 2 ×
150 bp setup at the Science for Life Laboratory (Stockholm) at the
SNP&SEQ Technology Platform over an entire Illumina NovaSeq6000
SP4 flow cell. The raw data are accessible at NCBI (SRA accession:
PRJNA627699). The bioinformatic procedure is described in detail in
the
3
RNA-seq and ChIP-seq Library Preparation
4
Transcriptomic Analysis of Smad Decoy Effects
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
HSFs were seeded onto a 12-well plate at 1×105 cells per well. Following a 24-h incubation, the cells were left untreated or transfected with 100 nM of the Smad decoy mixed with PepC (Pepscan). Following a 72-h transfection, the cells were harvested and the total RNA was isolated using DirectZol kit (Zymo Research Corp.). The barcoded mRNA sequencing library was subsequently prepared using TruSeq RNA Library Prep v2 (Illumina, Inc.), according to the manufacturer's protocol. The resultant libraries were pooled and sequenced on a HiSeq 2000 sequencer (Illumina, Inc.). Following removal of the adaptor sequences using Prinseq, the sequences were mapped to the human transcriptome (hg19 genome using TOPHAT (18 (link)) and the aligned BAM file was used to read counts for each gene using HTSeq (19 (link)). Subsequent analysis was performed using normalised counts and standard deviations derived from the 3-fold-changes of individual genes between the untreated and treated samples. Gene ontology analysis was performed by analysing the 178 most differentially regulated genes using the DAVID analysis algorithm v6.8 hosted on the website (https://david.ncifcrf.gov ) using the GOTERM_CC_DIRECT (cellular component) annotation (20 (link)).
5
Multiomics analysis of sediment microbiome
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
DNA and RNA were extracted from thawed and homogenized sediment with the DNeasy PowerMax soil kit (Qiagen) and RNeasy PowerSoil kit (Qiagen), with 10 g and 2 g input material, respectively. Extracted RNA was DNase treated with the Turbo DNA-free kit (Invitrogen) and rRNA depletion using the RiboMinus transcriptome isolation kit (bacterial version, ThermoFisher Scientific). Multiplexed libraries were prepared with the ThruPLEX DNA-seq (Rubicon Genomics) and TruSeq RNA Library Prep v2 [Illumina, without the poly(A) selection step] kits for DNA and RNA, respectively. Paired-end 2- by 150-bp sequencing was conducted at the Science for Life Laboratory, Stockholm, Sweden, on the Illumina NovaSeq platform (DNA on one lane, NovaSeq6000 S2, and RNA on one lane, Illumina NovaSeq6000 S4). The sequencing yielded on average 41 million (range 32 to 52) and 82 million (range 74 to 89) read-pairs for each DNA and RNA sample, respectively.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!