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Title: Next Generation Sequencing Facilitates Quantitative Analysis of Altered Expression of miRNAs and piRNAs in the Testes of Mtdh exon 3-deficient Mice      
dateReleased:
06-17-2015
description:
Purpose: To identify the correlation of the expression of testicular miRNAs and piRNAs with the infertility of Mtdh exon 3 deficient mice Methods: Total RNAs from individual testis of two WT mice and three homozygous Mtdh exon 3-depleted mice were used to prepare the miRNA sequencing library. After the completed libraries were quantified with an Agilent 2100 Bioanalyzer, the DNA fragments in the libraries were denatured with 0.1M NaOH to generate single-stranded DNA molecules, captured on Illumina flow cells, amplified in situ and sequenced for 36 cycles on Illumina HiSeq 2000 according to the manufacturer’s instructions. Image analysis and base calling were performed using Off-Line Basecaller software (OLB V1.8.0). Subsequently, 3’ adapter sequences were trimmed from clean reads (reads that passed Solexa CHASTITY quality filter) and any reads shorter than 15nt were discarded. Next the 3’-adapter-trimmed-reads (≥ 15nt) were aligned to the latest known human reference miRNA precursor set (Sanger miRBase 19) using Novoalign (v2.07.11). Reads (counts < 2) were discarded when calculating miRNA expression. In order to characterize the isomiR variability, any sequences that matched the miRNA precursors in the mature miRNA region ±4nt (no more than one mismatch) were accepted as mature miRNA isomiRs, which were grouped according to the 5-prime (5p) or 3-prime (3p) arm of the precursor hairpin. For piRNAs, the 3’-adapter-trimmed reads (length ≥ 15nt) were aligned to the latest piRNA set in piRNAbank using Novoalign software (v2.07.11). Expression of each piRNA was defined as the mapped tag counts. Results: Reduced expression of miRNAs was detected in homozygous Mtdh exon 3-deficient testes (Table I). As shown in tables II and III, piRNA expression levels were dysregulated, with some being reduced and others increased in homozygous Mtdh exon 3-deficient testes compared to WT mice. Conclusions: Our study represents the first detailed analysis of Mtdh deficiency on the expression of small noncoding RNAs, generated by RNA-seq technology. We conclude that Mtdh is correlated to the expression of small non-coding RNAs including miRNAs and piRNAs at testis of mouse. Examine expression of miRNAs and piRNAs at testes of wild type and Mtdh deficent mice
privacy:
not applicable
aggregation:
instance of dataset
ID:
E-GEOD-62330
refinement:
raw
alternateIdentifiers:
62330
keywords:
functional genomics
dateModified:
06-20-2015
availability:
available
types:
gene expression
name:
Mus musculus
accessURL: https://www.ebi.ac.uk/arrayexpress/files/E-GEOD-62330/E-GEOD-62330.raw.1.zip
storedIn:
ArrayExpress
qualifier:
gzip compressed
format:
TXT
accessType:
download
authentication:
none
authorization:
none
accessURL: https://www.ebi.ac.uk/arrayexpress/files/E-GEOD-62330/E-GEOD-62330.processed.1.zip
storedIn:
ArrayExpress
qualifier:
gzip compressed
format:
TXT
accessType:
download
authentication:
none
authorization:
none
accessURL: https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE62330
storedIn:
Gene Expression Omnibus
qualifier:
not compressed
format:
HTML
accessType:
landing page
primary:
true
authentication:
none
authorization:
none
abbreviation:
EBI
homePage: http://www.ebi.ac.uk/
ID:
SCR:004727
name:
European Bioinformatics Institute
homePage: https://www.ebi.ac.uk/arrayexpress/
ID:
SCR:002964
name:
ArrayExpress

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