PCB 3063 General Genetics Lecture Notes Final Exam new material only This set of notes is really long If you want to find notes for a specific date ex 1 27 or topic codominance press Ctrl F on your keyboard Command F for Mac users and type in what you want to find It ll search the document for whatever specific word phrase you typed in and save you the time of scrolling through everything Start Final Exam Mon 4 7 Eukaryotes Transcription in nucleus 3 polymerases RNA Pol II pre mRNA Cis acting elements Cis genes on same DNA strand Promoter upstream regulatory DNA sequences Core promoter Consensus sequences 5 TATA 3 at 30 base pairs before 1 base so negative bases GC region before TATA CAT box 100 bases 1 base initiator element flanks 1 site Certain downstream area 30 bases More specific consensus sequences for specific transcription factors Polymerase Works w general transcription factors TF II RNA Pol II There are dif types of TF II A B C etc Assemble at promoter is preinitiator complex Specificity regulated by specific transcription factors don t express all genes TF IID TBP TBP TATA Binding Protein Unzips helix Uses other TF II s RNA Pol II Specific proteins bind to specific DNA sequences or other proteins Immunoprecipitation method that shows which proteins bound to ea other Use antibodies to catch a protein Centrifuge it to see what else is attached to it PCR finds a specific sequence Another antibody used for 2nd protein Enhancers found upstream downstream or intergenic wedged inside DNA helix Can be 50 000 bases away from 1 start site Regulate expression Increase processivity of RNA Pol II complex Processing Pre mRNA transcription 25 000 genes 200 000 proteins because of dif transcription Pol II transcripts pre mRNA does post transcriptional processing RNA Pol II sits on unzipped transcription bubble Heterogenous nuclear RNA freshly copied RNA no modifications mRNA 5 end gets a 7 methyl guanicine cap 5 cap Cap 5 5 linkage triphosphate protect end from nuclease DNA eating enzyme 2 OH on base 1 and 2 methylated Additions happen during transcription Viruses mimic cap and methylation Polyadenylation happens after termination of transcription 3 end of most eukaryotic RNA goes thru this Consensus recognition sequence recognized by RNA endonuclease Endonuclease cuts piece from middle of strand Exonuclease cuts off end of strand RNA endonuclease cuts during transcription Poly A polymerase extends 3 OH end with string of A s Creates poly A tail prevents ends from decaying and info getting lost Exon part of DNA strand that s expressed kept Intron part of DNA strand that s intervening cut out Termination stop RNA Pol II ends transcription Rat I protein exonuclease Cleaves nucleotides from RNA downstream of cut site Falls off RNA Splicing 40 60 all human pre mRNA spliced Removes introns combines exons also remove exons Alters protein s being made Directed by consensus sequence in pre mRNA Snurps Small Nuclear Riboproteins Causes dif splicing to alter expression Lupus Erythematosus autoimmune disease Antibodies attacking snurps Spliceosome splices pre mRNA Forms on splice sites contains snurps Sn RNA Small Nuclear RNA Complimentary to intron consensus sequence Larial structure loop made of introns that were cut released Snurps connect end of exons introns Spliceosome helps break rebuild covalent bonds in neighbor nucleotides Cut 5 end on intron and 3 end on exon Join exons together release introns Tues 4 8 Promoters not included on RNA strand Pre mRNA remove introns add 5 cap add poly A tail mRNA Cis element on same strand Trans element transcription factors Enhancers can be far from promoter region DNA strand loops on itself to make regions meet Spliceosomes attach to RNA on either side of intron Curl it around in loop to bring both exons together Intron bit of strand loops around gets cut off Fall off RNA as larial loop structure Areas 5 untranslated region in DNA not copied onto RNA Promoter in DNA not copied onto RNA 5 cap protects 5 end of mRNA AAUAA consensus sequence Transcription start site 1 base Exon in pre mRNA mRNA codes for protein Intron in pre mRNA not included in mRNA 3 untranslated region in DNA not copied onto RNA Poly A tail protects 3 end of mRNA Wed 4 9 Ex Heterozygous albinism Aa curled fingernail Cc Homozygous for lactose intolerance ll of dif gamete genotypes Aa 2 Cc 2 ll 1 2 2 1 4 Ex Mice colors Yellow mouse x yellow mouse 6 yellow 3 black What type of inheritance 2 1 ratio not 1 2 1 Mendelian inheritance ratio Missing the first 1 Recessive lethal homozygous dominant fatal Ex Flower colors Pink x white all pink F1 pink x pink 9 pink 4 white 3 cyan What do numbers demonstrate Epistasis cyan phenotype was masked Hox genes Ultrahithorax intron 70 000 bases long 2 000 base intron expressed more often DMD 2 3 mil bases in gene Tied to muscular dystrophy Splicing process Essential components see picture a 5 splice site defines exon intron boundary splice donor b 3 splice site defines intron exon boundary splice acceptor Polypyramidine tract at 3 end of intron Branch point A provides 2 OH for larial structure formation Covalent bond in larial loop uses 2 OH In order U1 Snurp On exon 1 and part of intron On exon1 On intron U2 snurp On intron 1st base A C 70 of time 2nd base A 60 of time 3rd base G 80 of time 1st base G 100 2nd base U 100 3rd base A G 95 of time 1st base U 90 of time Next base A G Next base A right before 2 OH site 2 OH U2AF snurp Between U2 snurp and U2AF Polypyramidine Base 1 A 100 of time Next base G 100 Exon 2 1st base G Splicing steps Phosphodiester bond right after exon 1 broken leaves 3 OH after the G Intron bends back on itself Phosphate group right after cut site attaches to 2 OH after the base A Forms larial structure Intron breaks off both exons bound to each other by phosphate group Mutations Snurps bind to specific sequences define ends of exons U2AF snurp at start of exon exon U1 snurp at end of exon G is essential to bind U1 U1 bound more likely for U2AF to bind too Ex mutation at U1 site on exon 2 bases now G A No U1 binding not recognized by snurp Exon 2 mistaken for intron and cut out Exon skipping info was lost If intron not cut RNA retains nonsense info End codon in intron may be misread as end codon for gene Which would lead to losing info after that point Ex mutation within exon that looks like splice site Mutated site gets read as splice site exon s info after that point is cut out Affected exon is shorter Cryptic splice site
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