Analysis options for high-throughput sequencing in miRNA expression profiling

Stokowy, Tomasz; Eszlinger, Markus; Świerniak, Michał; Fujarewicz, Krzysztof; Jarząb, Barbara; Paschke, Ralf; Krohn, Knut

doi:10.1186/1756-0500-7-144

Cited by 74 publications

(68 citation statements)

References 44 publications

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“…While mapping to known microRNA precursor databases can be a faster and more direct method to quantify microRNA expression, mapping smRNA reads to the entire genome is emerging as a consensus standard procedure (Motameny et al 2010, Farazi et al 2012, Stokowy et al 2014. Genome mapping allows identification of novel smRNAs, and can effectively identify transcripts of RNA classes such as rRNA, tRNA, and mRNA.…”

Section: Introductionmentioning

confidence: 99%

Evaluation of microRNA alignment techniques

Ziemann

Kaspi²,

El‐Osta³

2016

RNA

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Genomic alignment of small RNA (smRNA) sequences such as microRNAs poses considerable challenges due to their short length (∼21 nucleotides [nt]) as well as the large size and complexity of plant and animal genomes. While several tools have been developed for high-throughput mapping of longer mRNA-seq reads (>30 nt), there are few that are specifically designed for mapping of smRNA reads including microRNAs. The accuracy of these mappers has not been systematically determined in the case of smRNA-seq. In addition, it is unknown whether these aligners accurately map smRNA reads containing sequence errors and polymorphisms. By using simulated read sets, we determine the alignment sensitivity and accuracy of 16 short-read mappers and quantify their robustness to mismatches, indels, and nontemplated nucleotide additions. These were explored in the context of a plant genome (Oryza sativa, ∼500 Mbp) and a mammalian genome (Homo sapiens, ∼3.1 Gbp). Analysis of simulated and real smRNA-seq data demonstrates that mapper selection impacts differential expression results and interpretation. These results will inform on best practice for smRNA mapping and enable more accurate smRNA detection and quantification of expression and RNA editing.

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Section: Introductionmentioning

confidence: 99%

Evaluation of microRNA alignment techniques

Ziemann

Kaspi²,

El‐Osta³

2016

RNA

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“…Sequencing of 100 bp was performed with an Illumina HighScan-SQ sequencer at the sequencing core facility of the IZKF Leipzig (Faculty of Medicine, University Leipzig) using version 3 chemistry and flow-cell according to the manufacturer's instructions. Demultiplexing of raw reads, adapter trimming and quality filtering was done as described (Stokowy et al 2014). Subsequent analysis was limited to those reads in which the 3 ′ end of the Hsp70 mRNA was flanked by the adaptor sequence.…”

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confidence: 99%

Oligoadenylation of 3′ decay intermediates promotes cytoplasmic mRNA degradation in Drosophila cells

Harnisch

Cuzic-Feltens

Dohm

et al. 2016

RNA

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Post-transcriptional 3′ end addition of nucleotides is important in a variety of RNA decay pathways. We have examined the 3 ′ end addition of nucleotides during the decay of the Hsp70 mRNA and a corresponding reporter RNA in Drosophila S2 cells by conventional sequencing of cDNAs obtained after mRNA circularization and by deep sequencing of dedicated libraries enriched for 3 ′ decay intermediates along the length of the mRNA. Approximately 5%-10% of 3 ′ decay intermediates carried nonencoded oligo(A) tails with a mean length of 2-3 nucleotides. RNAi experiments showed that the oligoadenylated RNA fragments were intermediates of exosomal decay and the noncanonical poly(A) polymerase Trf4-1 was mainly responsible for A addition. A hot spot of A addition corresponded to an intermediate of 3′ decay that accumulated upon inhibition of decapping, and knockdown of Trf4-1 increased the abundance of this intermediate, suggesting that oligoadenylation facilitates 3 ′ decay. Oligoadenylated 3 ′ decay intermediates were found in the cytoplasmic fraction in association with ribosomes, and fluorescence microscopy revealed a cytoplasmic localization of Trf4-1. Thus, oligoadenylation enhances exosomal mRNA degradation in the cytoplasm.

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“…The benefit of these techniques consist of scalability, simplicity, with rising in DNA polymerase activities and products, with a reduction of miscalculation prone, and even more efficiently practicable with the ultimate goal of achieving precise real-time products [91].…”

Section: Third-generation Sequencing Technologymentioning

confidence: 99%

High Throughput Sequencing Advances and Future Challenges

Pervaiz¹,

Lotfi²,

Haider³

et al. 2017

J Plant Biochem Physiol

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High throughput sequencing (HTS) technologies were developed into indispensable for genomic investigation and recent hottest topic for research in the field of genomics, which can generate over 100 times more data in comparison with the most complicated capillary sequencers. Recent advances and developments in HTS using next generation sequencing techniques have become essential in the studies of digital gene expression profiling, in epigenomics, genomics, and transcriptomics. These methodologies are dexterous of sequencing multiple DNA molecules in corresponding; facilitate hundreds of millions of DNA molecules to be sequenced within a short period of time. Though, the expenses and time period have been significantly reduced; the inaccurate profiles and boundaries of the new policy differ considerably from those of earlier reported sequencing techniques. The technical developments and decreasing cost of NGS (Next Generation Sequencing) technology have made RNA sequencing (RNA-seq) as a worldwide popular technique for gene expression projects. Various approaches have been done for the standardization of RNA sequencing data, which have been materialized in the reports, contradictory, both in the type of bias modification and in the statistical approach. On the other hand, as data persistently build up, there has been no apparent consensus on the proper normalization techniques to be used or the impact of chosen methods on the downstream analysis. In the present article, we mentioned the key features of HT-NGS like, Key HTS platforms and different sequencing applications, ethical limitation and future prospective.

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Analysis options for high-throughput sequencing in miRNA expression profiling

Cited by 74 publications

References 44 publications

Evaluation of microRNA alignment techniques

Evaluation of microRNA alignment techniques

Oligoadenylation of 3′ decay intermediates promotes cytoplasmic mRNA degradation in Drosophila cells

High Throughput Sequencing Advances and Future Challenges

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