Quantitative Bias in Illumina TruSeq and a Novel Post Amplification Barcoding Strategy for Multiplexed DNA and Small RNA Deep Sequencing

Nieuwerburgh, Filip Van; Soetaert, Sandra; Podshivalova, Katie; Wang, Eileen Ay-Lin; Schaffer, Lana; Deforce, Dieter; Salomon, Daniel R.; Head, Steven R.; Ordoukhanian, Phillip

doi:10.1371/journal.pone.0026969

Cited by 54 publications

(43 citation statements)

References 12 publications

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“…The TruSeq Small RNA Sample Preparation Kit introduces the indexing barcode during PCR amplification of the library after adapter ligation. This method significantly reduces sample bias over previous indexing/barcoding approaches where a barcode was ligated directly to the miRNA (Van Nieuwerburgh et al 2011). Researchers can barcode up to 48 samples and load as many as desired per lane of a sequencing flow cell, depending upon the amount of small RNA coverage required by the experiment.…”

Section: Introductionmentioning

confidence: 99%

Identification of extracellular miRNA in human cerebrospinal fluid by next-generation sequencing

Burgos

Javaherian

Bomprezzi

et al. 2013

RNA

179

174

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There has been a growing interest in using next-generation sequencing (NGS) to profile extracellular small RNAs from the blood and cerebrospinal fluid (CSF) of patients with neurological diseases, CNS tumors, or traumatic brain injury for biomarker discovery. Small sample volumes and samples with low RNA abundance create challenges for downstream small RNA sequencing assays. Plasma, serum, and CSF contain low amounts of total RNA, of which small RNAs make up a fraction. The purpose of this study was to maximize RNA isolation from RNA-limited samples and apply these methods to profile the miRNA in human CSF by small RNA deep sequencing. We systematically tested RNA isolation efficiency using ten commercially available kits and compared their performance on human plasma samples. We used RiboGreen to quantify total RNA yield and custom TaqMan assays to determine the efficiency of small RNA isolation for each of the kits. We significantly increased the recovery of small RNA by repeating the aqueous extraction during the phenol-chloroform purification in the top performing kits. We subsequently used the methods with the highest small RNA yield to purify RNA from CSF and serum samples from the same individual. We then prepared small RNA sequencing libraries using Illumina's TruSeq sample preparation kit and sequenced the samples on the HiSeq 2000. Not surprisingly, we found that the miRNA expression profile of CSF is substantially different from that of serum. To our knowledge, this is the first time that the small RNA fraction from CSF has been profiled using next-generation sequencing.

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Section: Introductionmentioning

confidence: 99%

Identification of extracellular miRNA in human cerebrospinal fluid by next-generation sequencing

Burgos

Javaherian

Bomprezzi

et al. 2013

RNA

179

174

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“…However, the library preparation methods used in NGS seem to have systematic biased representation of the miRNAs (Linsen et al 2009;Tian et al 2010;Hafner et al 2011;Van Nieuwerburgh et al 2011). These biases can be introduced during ligation, cDNA synthesis, and PCR amplification.…”

Section: Introductionmentioning

confidence: 99%

Differences in microRNA detection levels are technology and sequence dependent

Leshkowitz

Horn‐Saban

Parmet

et al. 2013

RNA

106

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Identification and quantification of small RNAs are challenging because of their short length, high sequence similarities within microRNA (miRNA) families, and the existence of miRNA isoforms and O-methyl 3 ′ modifications. In this study, the detection performance of three high-throughput commercial platforms, Agilent and Affymetrix microarrays and Illumina next-generation sequencing, was systematically and comprehensively compared. The ability to detect miRNAs was shown to depend strongly on the platform and on miRNA modifications and sequence. Using synthetic transcripts, including mature, precursor, and Omethyl-modified miRNAs spiked into human RNA, a large intensity variation in all spiked-in miRNAs and a reduced capacity in detecting O-methyl-modified miRNAs were observed between the tested platforms. In addition, endogenous human miRNA expression levels were assessed across the platforms. Detected miRNA expression levels were not consistent between platforms. Although biases in miRNA detection were previously described, here the end-point result, i.e., detection intensity, of these biases was investigated on multiple platforms in a controlled fashion. A detailed exploration of a large number of attributes, including base composition, sequence structure, and isoform miRNA attributes, suggests their impact on miRNA expression detection level. This study provides a basis for understanding the attributes that should be considered to adjust platform-dependent detection biases.

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“…Double ligated miRNAs are reverse transcribed using RT primer hybridized to the 3´ adapter, and in the next step, PCR amplifi cation is performed. Generation of miRNA library is recently signifi cantly improved and allows attaching of barcode directly to the sequence during PCR step (van Nieuwerburgh et al 2011). Consequently, the library is run on a gel, size selected, and sequenced.…”

Section: Sequencingmentioning

confidence: 99%

Progress in micro RNA focused research in endocrinology

Voglova

Bezakova

Herichová

2016

Endocrine Regulations

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Micro RNAs (miRNAs) are small regulatory molecules of increasing biologists' interest. miRNAs, unlikely mRNA, do not encode proteins. It is a class of small double stranded RNA molecules that via their seed sequence interact with mRNA and inhibit its expression. It has been estimated that 30% of human gene expression is regulated by miRNAs. One miRNA usually targets several mRNAs and one mRNA can be regulated by several miRNAs. miRNA biogenesis is realized by key enzymes, Drosha and Dicer. miRNA/mRNA interaction depends on binding to RNA-induced silencing complex. Today, complete commercially available methodical proposals for miRNA investigation are available. Th ere are techniques allowing the identifi cation of new miRNAs and new miRNA targets, validation of predicted targets, measurement of miRNAs and their precursor levels, and validation of physiological role of miRNAs under in vitro and in vivo conditions. miRNAs have been shown to infl uence gene expression in several endocrine glands, including pancreas, ovary, testes, hypothalamus, and pituitary.

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Quantitative Bias in Illumina TruSeq and a Novel Post Amplification Barcoding Strategy for Multiplexed DNA and Small RNA Deep Sequencing

Cited by 54 publications

References 12 publications

Identification of extracellular miRNA in human cerebrospinal fluid by next-generation sequencing

Identification of extracellular miRNA in human cerebrospinal fluid by next-generation sequencing

Differences in microRNA detection levels are technology and sequence dependent

Progress in micro RNA focused research in endocrinology

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