mRNA was purified from total RNA using poly-T oligo-attached magnetic beads (Life Technologies, USA) and transcribed to cDNA using random oligonucleotides and M-MuLV Reverse Transcriptase (RNase H−) (TaKaRa, Dalian, China). NEBNext adaptor oligonucleotides (Illumia, USA) were ligated to 3′ ends of cDNA fragments. Then, 200-bp cDNA fragments were purified using the AMPure XP beads system (Beckman Coulter, USA). Ten cycles of PCR amplifications were performed to enrich cDNA fragments using the NEB Universal PCR primer and Index primer (Illumia, USA). The PCR products were purified using the AMPure XP beads system and quantified using the Agilent Bioanalyzer 2100 system. Finally, the four-coded samples were clustered by a cBot Cluster Generation System using the TruSeq PE Cluster Kit v3-cBot-HS (Illumia, USA), and then sequenced on an Illumina Hiseq 2000 platform.
Nebnext adaptor oligonucleotides
Manufactured by Illumina
Sourced in United States
NEBNext adaptor oligonucleotides are short, single-stranded DNA fragments used in next-generation sequencing library preparation. These adaptors are designed to be ligated to DNA fragments, enabling amplification and sequencing of the target DNA.
Automatically generated - may contain errors
Lab products found in correlation
Hiseq 2000 platform, by Illumina (2 mentions)
Poly t oligo attached magnetic beads, by Thermo Fisher Scientific (2 mentions)
Neb universal pcr primer and index primer, by Illumina (2 mentions)
Truseq pe cluster kit v3 cbot hs, by Illumina (2 mentions)
Agilent bioanalyzer 2100 system, by Agilent Technologies (1 mentions)
Ampure xp beads system, by Beckman Coulter (1 mentions)
Ampurexp bead system, by Agilent Technologies (1 mentions)
Klenow exo, by Illumina (1 mentions)
2 protocols using nebnext adaptor oligonucleotides
1
mRNA-seq Library Preparation Protocol
2
RNA-seq library preparation and sequencing
Check if the same lab product or an alternative is used in the 5 most similar protocols
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mRNA was purified from total RNA using poly-T oligo-attached magnetic beads (Life Technologies, CA, USA) and transcribed to cDNA using random oligonucleotides with M-MuLV Rease Transcriptase (RNase H-) (TaKaRa) and DNA polymerase I and RNase H (TaKaRa). NEBNext adaptor oligonucleotides (Illumina) were ligated to the 3' ends of cDNA fragments adenylated with Klenow Exo- (Illumina). Approximate 200 bp cDNA fragments were purified using the AMPure XP bead system (Beckman Coulter, Beverly, USA). Ten cycles of PCR amplifications were conducted to enrich cDNA fragments ligated to the adaptors on both ends using the NEB Universal PCR primer and Index primer (Illumina). The PCR products were purified using the AMPure XP bead system and quantified using the Agilent high sensitivity DNA (Agilent Technologies, CA, USA) on the Agilent 2100 bioanalyzer system. Finally, the four-coded samples were clustered by the cBot cluster generation system using the TruSeq PE Cluster Kit v3-cBot-HS (Illumina) according to the user manual, and then sequenced using the 100 bp protocol on an Illumina Hiseq 2000 platform.
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