AGBT 2010 - Nicole Cloonan - The University of Queensland
Intro to Stem Cells
* hot topic - potential for cell generating therapies
* Self renewable
Looking at Extracellular space network.
* molecules that control cell-cell interactions (among others)
* defines the pluripotent status of the cell
* protein-protein interactions
(Muller et al, Nature 455:401-505)
* 6 transcripts per gene on average
* so how does this affect the plurinet
* have a pipelien... [too fast]
* done SET and PET.
* 80% of tags map, 194M 50mers, 114M 25mers
Tags that don't map:
* LincRNA, intergene, etc...
* alternate splicing
* works well if you know what the annotations are.
* with PET, you can build transcript models if you don't have them already - learn more about alt. splice
* can be used for novel exon discovery
Chip-Seq from Ku et al
* Extended Exons. 3' exon extensions can be very long.
[Why is this Chip-Seq?]
Do Virtual Northerns
* Size fractionations
* What you find is that most annotated genes have the right refseq predicted lengths.
* however, some are shorter, some are longer
* Frequency at which tags from a particular library match predicted (based on refseq) vs from RNA data... You do see that some have very different results.
RNA are translated...
* if no signal peptide, cytoplasmic (on free ribosomes)
* if has signal, then it's translated by ribosomes bound to membranes
* use sucrose gradient to separate the two populations
* do PET, (35/75bp reads)
* compare signals in both fractions - they come up well in the predicted fraction.
* membrane associated RNA have very different proportions of extension (mainly long 3' UTRs) than those in the cytoplasmic fraction
MiRNA biogenesis and mRNA interactions.
* use fractionation to test
* RISC associated with polysomes (which works with fractionation)
* complexes stay together through fractionation
* Long UTRS are enriched for mRNA binding sites
Back to Plurinet
* Complexity is incredibly increased with the extra products and miRNA
* PET allows you to reconstruct loci level complexity from RNAseq data
* Size fractionation is useful
* translation state RNAseq allow s the capture of mRNA and miRNA data from polyribosomes
* Transcriptional complexity impacts greatly on interactions.
Labels: AGBT 2010