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33 Cards in this Set
- Front
- Back
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genomics
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-the study of genes and their functions
- generally at whole genome level -studies all genes stimulaneously |
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genetics vs genomics
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- differ in scale and focus
genetics: analyzes one or a few genes at a time genomics: collects data on all genes in an organisms |
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forward genetics
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- traditional
: look at mutant phenotype → detemine gene |
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reverse genetics
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gene sequence → determine function
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why invest in genomics?
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1. identify genes involved in dieases
2. find genes to target for drugs 3. identify enzymes for industrial app 4. study evolution 5. develop "high throughput" sequencing and computational tech to advance science |
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genomics strategies for finding genes
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1. sequence clones in cDNA lib
2. genomic sequencing |
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sequence clones in cDNA
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- generate Expressed Sequence Tags
* every cDNA comes from a gene |
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genomic sequencing
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-sequence all chromosomal DNA from organism
-bioinformatics - not all sequences come from a gene - genes are widely scattered |
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bioinformatics
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- computers put the sequences together and identify likely genes
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Strategies for genome sequencing
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1. sequence ordered clones
2. whole genome sequencing AKA shotgun |
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whole genome sequencing AKA shotgun
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get entire genome → fragments → clone → sequence 10,000 clones → align sequences with super computers → analyze to find gene
- challenge bc of repeating sequences |
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Assembling sequences
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1. clone small overlapping fragments → sequence each
-look for overlap → determine original sequence 2. computer alignments |
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ordered clone method AKA clone by clone method
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1. get large clones
2. subclone genome 3. sequence subclones 4. reconstruct c'some from sequence of subclones |
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what to do with a genome sequence?
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1. find genes (base on similarity to known genes)
2. predict functional components of promoters 3. study f(x) of gene |
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what program do you use to identify genes from c'some sequence data?
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Blast Program
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Blast program - - webtool bioinformatics tool
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- if ESTs( cDNA) were sequenced = checks for similarity to c'some
- "new" genes can be predicted by similarity to known genes (even from other species) -used to predict f(x) of "new" protein |
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how many frames can be translated
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→ 3 frames
← 3 frames bc + strand not always coding - strand can be coding too |
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DNA → protein. how many frames are needed? why?
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3 different frames bc 4th is same as the 1st
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what are data bases searched for?
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1. genes encoding a certain type of protein
2. patterns in DNA: transcription factor binding sites/cis elements |
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why do we look for promoters?
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if genes are expressed at the same time = may bind to same TF = should have similar sequences in their promoters
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what is important to find cis elements?
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conserved sequence upsteam
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what do you base studying function of gene on?
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1. transcription pattern
2. effect of mutation |
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how do you study function of a gene based on transcription pattern?
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-northern blots
- fusing to reporter genes - microarrays |
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micro arrays
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1. use robot to spot DNA from genes (cDNA) onto glass side
- 2 dots for 1 probe 2. prepare labeled cDNA from total mRNA tissue of interest 3. hybridize to slide 4. hybridization level reflects abundance of mRNA in tissue |
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how do you prepare labeled cDNA from mRNA tissue of interest ?
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- use reverse transcriptase
- oligo dT primer - labelled bases |
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Gene knock outs = gene disruptions
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- inactivate clone genes by insertional/deletion mutagenesis using DNA mediate transformation
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homologous integration
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- db crossover during transformation
- frequent in yeast |
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Deep sequence analysis/RNAsequence
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-expresses/shows anything that is expressed
- more powerful than mircoarrays -finds new genes |
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TDNA
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-integrates into host genome @ any possible position
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what are ways to regulate gene expression?
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1. transcriptional
2. postranscriptional 3. translational 4. post translational |
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transcriptional regulation of gene expression
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- # if genes
- sequence of promoter @ RNA pol binding site - DNA binds sites to +/- regulators - RNA pol associated proteins - chromatin structures |
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posttranscriptional regulation of gene expression
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- capping, polyadenylation, splicing
- transportation to cytoplasm - RNA stability - ribosome binding - codon bias |
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post translational regulation of gene expression
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- protein stability
- post translational modifications |
