Cucumis melo
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA144407 | ENA |
REPOSITORIES: ENA
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Project description:DNA microarrays are two-dimensional arrangements of specific probes deposited on a substrate that have been widely used in gene expression analysis by measuring mRNA accumulation. The use of this type of microarrays involves the synthesis of cDNA, which has to be double stranded (ds) if the microarray probes are of the positive strand. We have used a custom-synthesized non-commercial NimbleGen microarray from melon to evaluate an alternative method of ds cDNA synthesis, which differs substantially in its economical cost relative to a widely recommended method. The results suggested that both methods produce cDNA representative of the melon transcriptome to a similar extent, indicating that the alternative technique provides a cheaper method of ds cDNA synthesis for microarray gene expression assays.
2012-03-07 | GSE30693 | GEO
Project description:DNA microarrays are two-dimensional arrangements of specific probes deposited on a substrate that have been widely used in gene expression analysis by measuring mRNA accumulation. The use of this type of microarrays involves the synthesis of cDNA, which has to be double stranded (ds) if the microarray probes are of the positive strand. We have used a custom-synthesized non-commercial NimbleGen microarray from melon to evaluate an alternative method of ds cDNA synthesis, which differs substantially in its economical cost relative to a widely recommended method. The results suggested that both methods produce cDNA representative of the melon transcriptome to a similar extent, indicating that the alternative technique provides a cheaper method of ds cDNA synthesis for microarray gene expression assays. Recently, we have analyzed the transcriptome of melon in response to WMV infection. Cotyledons of two genotypes of melon were virus inoculated and transcriptomic responses to the infection were analyzed by comparing infected and mock inoculated samples at 1, 3, and 7 days post-inoculation (dpi). Three biological replicates were performed for each sample. Double stranded cDNA was obtained with the Double stranded cDNA synthesis kit (Invitrogen, Carlsbad, CA, USA), based on the nick translation approach (Mol. Cell. Biol (1982) 2:161-170; Gene (1983) 25:263-269). Raw and processed microarray data are freely available from GEO database under the accession number GSE30111. By using this set of microarray hybridizations as a reference, RNA corresponding to infected cotyledons replicate 3 at 1 dpi (A1) and replicate 1 at 3 dpi (A2) (GEO accession numbers GSM745566 and GSM745567) were used to perform cDNA synthesis by the alternative method (samples B1 and B2, respectively), based on the SMART approach (BioTechniques (2001) 30:892-897), and microarray data were compared.
2012-03-07 | E-GEOD-30693 | biostudies-arrayexpress
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2018-12-28 | GSE124419 | GEO
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Project description:The tracks show enrichment of RNA sequence tags mapped to the mouse genome generated by high throughput sequencing (RNA-Seq). Double stranded cDNA was synthesized from enriched RNA that was obtained after depletion of ribosomal RNA. Pieces of cDNA, 300-350 nucleotides in length, were PCR amplified, adapter ligated, and sequenced on an Illumina HiSeq sequencer. For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf Cells were grown according to the approved ENCODE cell culture protocols (http://hgwdev.cse.ucsc.edu/ENCODE/protocols/cell). Total RNA was extracted using RNeasy Mini Kit (74104, Aiagen), following the manufacturer's protocol. Ribosomal RNA was removed from total RNA using the Ribo-ZeroTM Gold Kits (MRZG126, Epicentre). Double-stranded cDNA synthesis was performed on the rRNA depleted RNA using random primers and the SuperScript double-stranded cDNA synthesis kit (11917-010, Life Tech). After first strand cDNA synthesis, NucAway Spin Column (Ambion cat. 100070-30) was used to remove dNTPs. In the second strand cDNA synthesis reaction, dTTP in the dNTP mix was substituted with dUTP. After end repair and addition of 'A' base to 3' end, illumina paired-end adapter was ligated to Double-stranded cDNA library. After gel size selection of adapter ligated cDNA (300-350), Uracil-N-Glycosylase (UNG: Applied Biosystems) was used to digest the second strand cDNA (Parkhomchuk et al. , 2009). PCR amplified adapter ligated cDNA was sequenced using Illumina HiSeq. Sequence reads of 2x101 nt long with 0-2 mismatches were mapped to the mouse genome (version mm9) using the BWA aligner, version 0.5.7. The signal height corresponds to the number of overlapping fragments at each nucleotide position in the genome.
2012-07-24 | E-GEOD-39619 | biostudies-arrayexpress
Project description:Investigation of genome-wide expression in the mutant of histone H3K9 methyltransferase KRYPTONITE (KYP) or DNA methyltransferase CHROMOMETHYLASE3 (CMT3) in Arabidopsis. These mutants showed decrease in H3K9 methylation and DNA methylation levels, and transcriptional activation at transposons and repeats. Using NimbleGen DNA microarray, global pattern of expression of genes and transposons were examined in these mutants. Total RNA from leaves was isolated using the RNeasy Plant Mini kit (Qiagen) and was treated with DNase I (TAKARA). Double-stranded cDNA was synthesized using the SuperScript Double Stranded cDNA Synthesis kit (Invitrogen) and oligo (dT)20 primer. cDNA was labeled by Cy3, hybridized to 4 x 72k array, and scanned according to manufacture's instructions (www.nimblegen.com).
2010-09-21 | E-GEOD-22957 | biostudies-arrayexpress