Non-coding RNA tiling array in rice
Ontology highlight
ABSTRACT:
ORGANISM(S): Oryza sativa
PROVIDER: GSE6922 | GEO | 2007/02/06
SECONDARY ACCESSION(S): PRJNA98141
REPOSITORIES: GEO
Similar Datasets
Project description:The 1 Mb tiling array was used to represent the 10.0 Mb to 11.0 Mb region of japonica rice Chromosome 10. A tile path was designed with 36-mer probes at a step of 5 nt. All 388,994 probes were retained and synthesized into a single array. This array was hybridized to the cDNA target used for the full-genome tiling array as well as targets prepared from non-coding RNA sample I and II. Keywords: noncoding RNA Non-coding RNA species were isolated from total RNA as previously described (5). Briefly, total RNA isolated from japonica rice leaves using the Trizol method was loaded onto an Qiagen-tip and eluted with 0.6~1.0M NaCl gradient of QRW2 buffer (Qiagen) at 50 oC. Fractions corresponding to tRNAs (sample I) and small RNAs of 80-500 nucleotides (nt) (sample II) were collected. To remove remaining mRNAs and rRNAs, the small RNA fractions were hybridized to a mixture of specifically designed oligos in the binding buffer. Unwanted RNA molecules were targeted and removed by a magnetic bead based process (MicrobExpress kit, Ambion). The enriched non-coding RNA pool was then dephosphorylated with a calf intestine alkaline phosphatase and ligated to a 3ï¢ adaptor oligonucleotide by the T4 RNA ligase (5). Small size molecules and excessive adaptors were removed from the ligation products using the mirVana miRNA isolation kit (Ambion) and reverse transcribed at 50 oC using the Thermoscript (Invitrogen) and the oligo 3RT as the primer (5). Amino-allyl-modified dCTP was incorporated into the cDNA strand during reverse transcription.
2010-06-11 | E-GEOD-6922 | biostudies-arrayexpress
Project description:In the study of gene expression regulation in plants, it is often desirable to distinguish transcript pools derived from different alleles present in the same organism. We report here an oligonucleotide tiling microarray designed to specifically target 518 single nucleotide polymorphisms (SNPs) between the two sequenced rice (Oryza sativa) subspecies indica and japonica. The tiling array includes all pairs of 25-mer allele-specific probes interrogating each SNP by placing the polymorphic site at all 25 possible positions within the probe. Hybridization of the tiling array to a titration series in which the japonica- and indica-derived cDNA templates are mixed with altering proportions and development of a novel bioinformatic methodology allowed us to screen for diagnostic probe pairs for each SNP. Our result indicates that 284 (55%) SNPs have at least one diagnostic probe pair suitable for distinguishing and quantifying the relative abundance of allele-specific transcripts. As a proof-of-concept, we analyzed allele-specific expression in reciprocal indica × japonica F1 hybrids and detected imbalanced expression at 95 (33%) and 71 (25%) SNPs, respectively. Comparison results from RNA-sequencing and allele-specific real-time PCR experiment validates the sensitivity and reliability of the tiling array method. Together, our results demonstrate the advantages of the tiling array method in interrogating large numbers of SNPs and for the reliability of the experimental and analytical techniques used for quantifying allele-specific gene expression.
2010-07-01 | GSE20678 | GEO
Project description:In the study of gene expression regulation in plants, it is often desirable to distinguish transcript pools derived from different alleles present in the same organism. We report here an oligonucleotide tiling microarray designed to specifically target 518 single nucleotide polymorphisms (SNPs) between the two sequenced rice (Oryza sativa) subspecies indica and japonica. The tiling array includes all pairs of 25-mer allele-specific probes interrogating each SNP by placing the polymorphic site at all 25 possible positions within the probe. Hybridization of the tiling array to a titration series in which the japonica- and indica-derived cDNA templates are mixed with altering proportions and development of a novel bioinformatic methodology allowed us to screen for diagnostic probe pairs for each SNP. Our result indicates that 284 (55%) SNPs have at least one diagnostic probe pair suitable for distinguishing and quantifying the relative abundance of allele-specific transcripts. As a proof-of-concept, we analyzed allele-specific expression in reciprocal indica × japonica F1 hybrids and detected imbalanced expression at 95 (33%) and 71 (25%) SNPs, respectively. Comparison results from RNA-sequencing and allele-specific real-time PCR experiment validates the sensitivity and reliability of the tiling array method. Together, our results demonstrate the advantages of the tiling array method in interrogating large numbers of SNPs and for the reliability of the experimental and analytical techniques used for quantifying allele-specific gene expression. For each selected SNP, 25 pairs of 25-mer oligonucleotide probes were designed with one match perfectly to the indica allele and the other the japonica allele within each pair. Among the 25 probe pairs, the polymorphic site was placed at a different position such that all possible 25-mer allele-specific probes spanning the polymorphic site were included. The resultant 25,900 probes were synthesized in triplicate in a single microarray produced on the Maskless Array Synthesizer platform as previously described (Li et al., 2005; 2006). Poly(A+) RNA from japonica, indica and their reciprocal hybrids was reverse transcribed using an oligo(dT)18 primer, during which amino-allyl-modified dUTP (aa-dUTP) was incorporated. The aa-dUTP decorated cDNA was fluorescent labeled by conjugating the monofunctional Cy3 dye (GE Healthcare) to the amino-allyl functional groups in the cDNA. Dye-labeled target was quantified using a spectrophotometer and two μg used for hybridization to each array. In the titration series experiment, the dye-labeled targets from japonica and indica were mixed in known proportions before hybridization.
2010-07-01 | E-GEOD-20678 | biostudies-arrayexpress
Project description:This experiment was designed to identify transcribed regions of japonica subspecies of the rice genome. A series of high-density oligonucleotide tiling arrays that represent sense and antisense strands of the entire nonrepetitive sequence of all the 12 chromosomes were designed to measure genome-wide transcription. A total of 12253842 36mer oligonucleotide probes positioned every 46 nt on average were used for this purpose. The probes were synthesized via maskless photolithography at a feature density of approximately 389,000 probes per slide. The arrays were hybridized with fluorescence-labeled cDNA reverse-transcribed from equal amounts of four selected poly(A)+ RNA population (seedling root, seedling shoot, panicle, and suspension cultured cells). Keywords: tiling array, genome-wide transcription
2007-02-21 | GSE6996 | GEO
Project description:Experimentally mapped transcriptome structure of Sulfolobus solfataricus P2 by hybridizing total RNA (including RNA species <200 nt) to genome-wide high-density tiling arrays (60 mer probes tiled every 25 nt).
2011-06-27 | GSE26779 | GEO
Project description:Experimentally mapped transcriptome structure of Methanococcus maripaludis S2 by hybridizing total RNA (including RNA species <200 nt) to genome-wide high-density tiling arrays (60 mer probes tiled every 14 nt).
2011-06-27 | GSE26777 | GEO
Project description:Experimentally mapped transcriptome structure of Pyrococcus furiosus DSM 3638 by hybridizing total RNA (including RNA species <200 nt) to genome-wide high-density tiling arrays (60 mer probes tiled every 16 nt).
2011-06-27 | GSE26778 | GEO
Project description:Experimentally mapped transcriptome structure of Escherichia coli BL21(DE3) by hybridizing total RNA (including RNA species <200 nt) to genome-wide high-density tiling arrays (60 mer probes tiled every 10 nt).
2017-01-13 | GSE93565 | GEO
Project description:This experiment was designed to identify transcribed regions of both japonica and indica rice chromosome 10. A series of high-density oligonucleotide tiling arrays that represent sense and antisense strands of the entire nonrepetitive sequence of the chromosome were used to measure transcriptional activities. A total of 750,282 and 838,816 36mer oligonucleotide probes, positioned every 46 nt on average, were designed to interrogating the japonica and the indica chromosome, respectively. The probes were synthesized via maskless photolithography at a feature density of approximately 389,000 probes per slide. The arrays were hybridized with fluorescence-labeled cDNA reverse-transcribed from equal amounts of four selected poly(A)+ RNA populations, namely, seedling roots, seedling shoots, panicles, and suspension cultured cells of the respective rice subspecies. Keywords: other
2005-04-09 | GSE2500 | GEO
Project description:Heat-responsive and time-resolved changes in transcriptome of E. coli BL21(DE3) Experimentally mapped transcriptome structure of Escherichia coli BL21(DE3) by hybridizing total RNA (including RNA species <200 nt) to genome-wide high-density tiling arrays (60 mer probes tiled every 10 nt).
2020-04-12 | GSE148034 | GEO