Small RNA sequencing identifies miRNA roles in ovule and fiber development
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ABSTRACT:
ORGANISM(S): Gossypium hirsutum
PROVIDER: GSE60380 | GEO | 2014/08/16
SECONDARY ACCESSION(S): PRJNA258280
REPOSITORIES: GEO
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Project description:To dissect the roles of miRNAs in fiber development, we sequenced small RNAs from ovules -1 to +1 day post anthesis (DPA) and young leaves. A series of conserved and novel miRNA were identified from cotton EST database and the genome of G.raimondii, many of which were shown to be expressed differentially between ovule and leaf, indicating their potential roles in fiber development or leaf development. Cotton (Gossypium Hirsutum L.) cultivar TM-1 were grown in the greenhouse. Cotton ovules -1 to +1 day post anthesis (DPA) and young leaves were harvested for five biological replicates and immediately frozen in liquid nitrogen. They were then stored at -80M-BM-0C following RNA extraction. Totals RNAs were extracted from each tissue sample using the mirVana miRNA isolation kit (Ambion, Austin, TX) according to the manufacturerM-bM-^@M-^Ys protocol. The small RNA samples extracted from the five biological replicates were pooled together for leaf and ovule, respectively. Finally, the construction of pooled small RNA libraries and sequenced were performed by LC Sciences (Houston, TX) using Illumina high-throughput sequencing platform.
2014-08-16 | E-GEOD-60380 | biostudies-arrayexpress
Project description:RNAs from the upland cotton 9-DPA fibers were compared to the 9-DPA fiber-detached ovule. RNAs from the upland cotton 9-DPA fibers were compared to the 9-DPA fiber-detached ovule.
2010-07-01 | E-GEOD-4642 | biostudies-arrayexpress
Project description:Three samples of cotton fiber harvested 3, 7 and 15 days post anthesis (DPA) and a sample of cotton ovule harvestd 0 DPA were used for non-coding RNA extraction and seloxa sequencing experiments, respectively.Cotton plants (Gossypium hirsutum cv. Xuzhou 142) were grown in a soil mixture in fully automated walk-in growth rooms with 300 M-BM-5mol mM-bM-^@M-^S2 sM-bM-^@M-^S1 average light intensity, 60% relative humidity and temperatures set to 30C during the light period and 28C during the dark (12-h light/dark cycle). These conditions were consistent throughout the year. Small RNA molecules under 30 bases were amplified and isolated from an agarose gel. The purified DNA was used directly for cluster generation and sequencing analysis using the Illumina Genome Analyzer IIx according to the manufacturer's instructions. The 38nt sequence tags from sequencing went through data cleaning first, which included getting rid of the low-quality tags and clipping adapter sequences. Three samples of cotton fiber harvested 3, 7 and 15 days post anthesis (DPA) and a sample of cotton ovule harvestd 0 DPA were used for non-coding RNA extraction and seloxa sequencing experiments, respectively.
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Project description:RNAs from the upland cotton 9-DPA fibers were compared to the 9-DPA fiber-detached ovule. Keywords: tissue comparison
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Project description:Cotton is one of the most commercially important Fiber crops in the world and used as a source for natural textile Fiber and cottonseed oil. The fuzzless-lintless ovules of cotton mutants are ideal source for identifying genes involved in Fiber development by comparing with Fiber bearing ovules of wild-type. To decipher molecular mechanisms involved in Fiber cell development, transcriptome analysis has been carried out by comparing G. hirsutum cv. MCU5 (wild-type) with its fuzzless-lintless mutant (MUT). Cotton bolls were collected at Fiber initiation (0 dpa/days post anthesis), elongation (5, 10 and 15 dpa) and secondary cell wall synthesis stage (20 dpa) and gene expression profiles were analyzed in wild-type and MUT using Affymetrix cotton GeneChip Genome array.
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Project description:Cotton fibers are seed trichomes, and their development undergoes a series of rapid and dynamic changes from fiber cell initiation, elongation to primary and secondary wall biosynthesis and fiber maturation. Previous studies showed that cotton homologues encoding putative MYB transcription factors and phytohormone responsive factors were induced during early stages of ovule and fiber development. Many of these factors are targets of microRNAs (miRNAs). miRNAs are ~21 nucleotide (nt) RNA molecules derived from non-coding endogenous genes and mediate target regulation by mRNA degradation or translational repression. Here we show that among ~4-million reads of small RNAs derived from the fiber and non-fiber tissues, the 24-nt small RNAs were most abundant and were highly enriched in ovules and fiber-bearing ovules relative to leaves. A total of 28 putative miRNAs families, including 25 conserved and 3 novel miRNAs were identified in at least one of the cotton tissues examined. Thirty-two pre-miRNA hairpins representing 19 unique families were detected in Cotton Gene Indices version 9 (CGI9) using mirCheck. Sequencing, miRNA microarray, and small RNA blot analyses showed that many of these miRNAs differentially accumulated during ovule and fiber development. The cotton miRNAs examined triggered target cleavage in the same predicted sites of the cotton targets in ovules and fibers as that of the orthologous target genes in Arabidopsis. Targets of the potential new cotton miRNAs matched the previously characterized ESTs derived from cotton ovules and fibers. The miRNA targets including those encoding auxin response factors were differentially expressed during fiber development. We suggest that both conserved and new miRNAs play an important role in the rapid and dynamic process of fiber and ovule development in cotton.
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Project description:DNA methylation is essential for plant and animal development. In plants, methylation occurs at CG, CHG, and CHH (H = A, C or T) sites via distinct pathways. Cotton is an allotetraploid consisting of two progenitor genomes. Each cotton fiber is a rapidly-elongating cell derived from the ovule epidermis, but the molecular basis for this developmental transition is unknown. Here we analyzed methylome, transcriptome, and small RNAome and revealed distinct changes in CHH methylation during ovule and fiber development. In ovules, CHH hypermethylation in promoters correlated positively with siRNAs, inducing RNA-dependent DNA methylation (RdDM), and up-regulation of ovule-preferred genes. In fibers, the ovule-derived cells generated additional heterochromatic CHH hypermethylation independent of RdDM, which repressed transposable elements (TEs) and nearby genes including fiber-related genes. Furthermore, CHG and CHH methylation in genic regions contributed to homoeolog expression bias in ovules and fibers. Inhibiting DNA methylation using 5-aza-2'-deoxycytidine in cultured ovules has reduced fiber cell number and length, suggesting a potential role for DNA methylation in fiber development. Thus, RdDM-dependent methylation in promoters and RdDM-independent methylation in TEs and nearby genes could act as a double-lock feedback mechanism to mediate gene and TE expression, potentiating the transition from epidermal to fiber cells during ovule and seed development.
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Project description:This experiment was designed to investigate the molecular basis of cotton fiber cell initiation. 32,000 ESTs were sequenced from Gossypium hirsutum L. TM-1 immature ovules (GH_TMO) and developed cotton oligonucleotide microarrays containing ~23,000 unigenes. Transcriptome analyses were performed to compare gene expression changes in laser capture microdissected fiber cell initials (or epidermis) and inner ovules. The gene expression profiles of the fiber cell initials were compared with those of the inner ovules in each developmental stage prior to, right at, and shortly after the initiation of fiber cells. Many genes in various molecular function or biological processes were over- or under-represented between fibers and non-fiber tissues in each developmental stage, suggesting temporal regulation of gene expression during early stages of fiber development. For gene expression studies using a large set cotton oligo-microarray, 4 developmental stages were chosen. To study differential expression during fiber initiation, ovules at -2 DPA, 0 DPA, and 2 DPA were used. One of the fiber elongation stage tissues (7 DPA) was included. In each developmental stage, epidermis was separated from inner ovules and subjected to the hybridization. In addition, epidermis and ovule comparisons were performed individually with 0 DPA as a control point for comparison.
2011-11-02 | E-GEOD-17378 | biostudies-arrayexpress
Project description:The chip of cotton oligonucleotide microarrrays, which contain ~23,000 UniGenes from our own ESTs sequence project (Gossypium hirsutum L. TM-1 immature ovules (GH_TMO) ) and open resources, was developed. Transcriptome analyses were performed to compare gene expression changes in laser capture microdissected (LCM) fiber cell initials (or epidermis) and inner ovules. The gene expression profiles of the fiber cell initials were compared with those of the inner ovules in each developmental stage prior to (-2DPA), right at (0DPA), and shortly after the initiation of fiber cells (2DAP and 7DPA). Many genes in various molecular functions or biological processes were over- or under-represented between fibers and non-fiber tissues in each developmental stage, suggesting temporal regulation of gene expression during early stages of fiber development. For gene expression studies using a large set cotton oligo-microarray, 4 developmental stages were chosen. To study differential expression during fiber initiation, ovules at -2 DPA, 0 DPA, and 2 DPA were used. One of the fiber elongation stage tissues (7 DPA) was included. In each developmental stage, epidermis was separated from inner ovules and subjected to the hybridization. In addition, epidermis and ovule comparisons were performed individually with 0 DPA as a control point for comparison.
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