transcriptome what is it - genome wide transcript abundance
DESCRIPTION
Transcriptome What is it - genome wide transcript abundance How do you obtain it - Arrays + MPSS What do you do with it when you have it - ?. Affymetrix GeneChip technology. Quality control steps. 1. RNA quality 260/280 ratio. 2. Array image inspection. 3. Hybridization controls. - PowerPoint PPT PresentationTRANSCRIPT
Transcriptome
What is it - genome wide transcript abundance
How do you obtain it - Arrays + MPSS
What do you do with it when you have it - ?
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Affymetrix GeneChip technology
Quality control steps
1. RNA quality 260/280 ratio
2. Array image inspection
3. Hybridization controls
4. % genes present
5. Scaling and normalization factors
Affymetrix oligo array
Types of ArraysGene Expression Arrays
As name suggests for measuring expressionMost genes in Genome coveredFrom a variety of organisms
Probe sets are designed to 3’ end of geneas labelling of probe starts at “end” of gene
Exon ArraySimilar to gene expression arrayDiffers in that all exons are covered inprobe sets
Identifies alternative splicing
Tiling arraysCover Genome sequenceFor mapping - not for expressionCan map a single mutation in a single hydridisationChiP on Chip analysis
ChIP-on-CHIPIdentify transcription factor
Generate antibody
Isolate nuclei
Immuno precipitate antibody with isolated genomic DNA
Antibody will bind to transcription factor bound to nuclear DNA
Isolate and amplify antibody bound DNAHybridise onto oligo chip
Signal where DNA is bound
Identify set of genes regulated by a specific transcription factor
SNP ChipFor detecting PolymorphismsCan detect 50 to 100 thousand on a single chipCan be used to screen for unknown associations
duplicated spots in each block
2 x 10k Arabidopsis cDNA array
ADNc
Hybridation
laser 2laser 1
analyse
RT
Cy3 Cy5
ARN polyA+
Micro-arrays
http://mpss.udel.edu/tutorial/flash/mpss.swf