BINF 511 Lecture 4: BINF 511 Lecture 4A 31Jan2018
25 Jun 2018
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Lecture 4A: Genomic strategies and data
January 31, 2018
Sequencing strategies
One gene of interest at a time - how long will this take?
Whole genome sequencing - expensive!
oShotgun approach
Random large fragments
Ends of clones are sequenced
Contigs assembled from sequences
Sequence depth (6-10x coverage)
You want to go over the same sequence at least 6 times, we want to
make sure that what we get is accurate
oClone contig approach
Random sequences ordered by overlaps into a contig (contig = contiguous
sequence)
Clones screened and ordered
Sequenced selected BAC clones ("BAC-end sequencing")
150-200 kb
Fragments overlap because of partial digest
Chromosome walking
Sequence, design primer to this sequence…
Sequencing the transcriptome - only sequence the expressed genes, but not the intergenic
regions; we can know which genes are expressed
Genomic libraries: represents all DNA in an organism
oWhen people talk about a genomic library, they're talking about physical clones in the
tube/lab; not the sequences of genes - this is important for the midterm
Maps
Genetic [linkage] maps are important, but we won't address them much in this class
oObserved traits connected to markers
oWhen complete enough, genetic linkage groups are built
oEx: soybean linkage groups -> boiled down to 20 linkage groups
If done well enough, the number of linkage groups = the number of
chromosomes
Physical map
oClones sequenced, identified and ordered
oObtained by markers, probe labelling and sequencing
Expression data
ESTs = expressed sequence tags (this is a sequencing based technique)
oExtremely important before genome sequencing became affordable
oA short sequence read of a gene; not the whole gene sequence, just a 'tag' for it
oLarge scale EST sequencing; useful when genome is too large for budget
oDNA -> RNA -> cDNA
mRNA -> cDNA -> cDNA ligated to plasmids
How to extract mRNA?
find more resources at oneclass.com
find more resources at oneclass.com
Document Summary
You want to go over the same sequence at least 6 times, we want to make sure that what we get is accurate o. Random sequences ordered by overlaps into a contig (contig = contiguous sequence) Sequencing the transcriptome - only sequence the expressed genes, but not the intergenic regions; we can know which genes are expressed. Genomic libraries: represents all dna in an organism: when people talk about a genomic library, they"re talking about physical clones in the tube/lab; not the sequences of genes - this is important for the midterm. Genetic [linkage] maps are important, but we won"t address them much in this class: observed traits connected to markers, when complete enough, genetic linkage groups are built o. Ex: soybean linkage groups -> boiled down to 20 linkage groups. If done well enough, the number of linkage groups = the number of chromosomes. Clones sequenced, identified and ordered: obtained by markers, probe labelling and sequencing.
