Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

RNA sequencing of chicken limb distal mesenchyme cells to compare gene expression over time and in heterochronic grafted cells

ABSTRACT: Distal limb mesenchyme cells were collected from embryonic chickens at Hamburger Hamilton (HH) stages HH24, HH27, and HH29, and also from samples that had been grafted from HH27 to HH20 and left to develop for 24 hours (equivalent to HH24). Samples were collected in triplicate (with 10 distal tips pooled per replicate), the RNA extracted, and sequenced on a HiSeq 2000 (Instrument: ST300). Bioinformatic and clustering analyses were performed to determine whether grafted cells are more similar to HH29 or HH24 samples, ie. which genes can be reset to expression levels observed in an earlier developmental stage.

INSTRUMENT(S): Illumina HiSeq 2000

ORGANISM(S): Gallus gallus

SUBMITTER: Matthew Towers

PROVIDER: E-MTAB-6437 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

An intrinsic cell cycle timer terminates limb bud outgrowth.

Pickering Joseph J Rich Constance A CA Stainton Holly H Aceituno Cristina C Chinnaiya Kavitha K Chinnaiya Kavitha K Saiz-Lopez Patricia P Ros Marian A MA Towers Matthew M

eLife 20180903

The longstanding view of how proliferative outgrowth terminates following the patterning phase of limb development involves the breakdown of reciprocal extrinsic signalling between the distal mesenchyme and the overlying epithelium (e-m signalling). However, by grafting distal mesenchyme cells from late stage chick wing buds to the epithelial environment of younger wing buds, we show that this mechanism is not required. RNA sequencing reveals that distal mesenchyme cells complete proliferative o ...[more]

PMID: 30175958

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Project description:Comparison of global gene expression in the proximal and distal colonic epithelium in azoxymethane treated rats. Experiment Overall Design: Ten male Sprague Dawley rats weighing approximately 245 Â± 14.5 g were housed in wire-bottomed caging in a temperature controlled room (22-24Â°C) with a 12 h light/dark cycle. They were randomly allocated into two groups (n=5) with approximately equal body weights. They were given free access to water and modified AIN-93G diet for 10 days when they were injected with 15 mg of AOM/kg subcutaneously (Sigma Chemical Co., St. Louis, MO, USA). They were anaesthetised with isoflurane and killed by exsanguination six hours after injection. The large bowel (excluding the rectum) was removed, opened longitudinally along the mesenteric border and digesta removed. The colon was rinsed clean with PBS and transferred to a chilled ceramic plate. The most distal and the most proximal 0.5 cm were discarded. Mucosal samples for gene expression and protein analyses were collected by scraping the next 4 cm of proximal and distal colon with new microscope slides. Experiment Overall Design: The mucosal scraping was transferred into RNAlater (Sigma Chemical Co., St. Louis, MO, USA) and stored at -80oC for later processing. All instruments were replaced or cleaned thoroughly between animals. All procedures involving animals were approved by the Commonwealth Scientific and Industrial Research Organisation (CSIRO) Human Nutrition Animal Ethics Committee and complied with the Australian code of practice (2004). Experiment Overall Design: The distal and proximal colonic mucosal samples were removed from the RNAlater stabilisation reagent (Sigma, Australia) and placed in 1ml of TRIzolÂ® Reagent (Invitrogen, Sydney, N.S.W., Australia). Samples were then homogenised using beads (mix of 2.5mm glass and 0.1 - 1.0mm diameter silicon-zirconian beads) in a MiniBeadbeater-8â¢ (BioSpec Products Inc. Oklahoma, US). Total RNA was then extracted according to the TRIzolÂ® Reagent manufacturerâs instruction after which samples were further purified using RNAeasy mini spin columns (QIAGEN, Doncaster, Victoria, Australia) with a DNase on-column digestion as per the manufactureâs instructions. The integrity of the RNA was checked using a Bioanalyzer 2100 (Agilent Technologies) and quantified using a NanoDropÂ® ND-1000 Spectrophotometer. Experiment Overall Design: RNA (4.5ug) samples were processed for Microarray expression analysis using high-density oligonucleotide arrays (AffymetrixÂ® GeneChip array, AffymetrixÂ®, Santa Clara, CA, USA) commensurate with the manufacturerâs instructions.

Dataset Information

RNA sequencing of chicken limb distal mesenchyme cells to compare gene expression over time and in heterochronic grafted cells

Publications

An intrinsic cell cycle timer terminates limb bud outgrowth.

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