Lecture #1 (Genome organization) Flashcards

Question

Bacetrial Conjugation

Answer 1

Bacteria form structure with another bacteria and exchnage information Usually exchnage a plasmid

Answer 2

Horizontal gene transfer is NOT only in bacteria Ex. Monsanto company makes genetic modified crops that will survive in the prescence of a chemical they made (chemical kills everything else) - Have a gene that allows the crop to survive in the presence of the chemical Farmers plant crops and spray chemical --> the crops survive and eveyrthing else dies BUT eventually the weeds came back - Weeds aquired the mansantos engineered gene = Horizontal gene transfer occured in plants

Answer 3

HGT may occur in mammales Don't REALLY have evidence BUT do have evidnece in H.pylori (causes ulcers) -- found that some genes in H/pylori are human in origin - Human genes went to H.pylori

Answer 4

Genomes of Eukryotic organeels are decended from bacteria (mitocndiral + chrlorplast genomes) - Chloroplast + mitocondra = look like bacteria (orginate from bacteria) Notable features: 1 ~10 genomes per mitocondria 2. ~8000 genomes per cell 3. Different genetic code 4. 37 genes 5. Proteins are all part of the electron transport chain

Answer 5

Prokayotes are comprised of bacteria and Archea Genome size is proportional to the number of genes Genes are frequencetey arranged as operons Have a capcity for rapid evolution through horizontal gene transfer Genomes of eukryotic organnelles are descended from free-living bacteria Gene density varies (Ex. yeast are more dense)

Answer 6

Always liner --> allows it to be bigger Chromosome number varies between difefrent Eukryotic - There is no corelation between chromosome number and complexity OR between gene number and complexity DNA is packaged Genes are discontinous gene density varies greatly

Answer 7

DNA is packaged - exists with nucleosomes Nucleosomes = protein complexes that wind DNA around it

Answer 8

Eukarotic genome has discontinous genes --> inteupted by introns - Have non-coding DNA within a gene AND the genes are far away from each other Most of mammalian genome is non-coding (90% doesn't code for a protein)

Answer 9

1990 - starting sequecning the human genome (all done by hand) Early 1990s - Sequenced yeast (test case for the human genome) - 1996 - yeast genome was finsihed sequenced (6,000 genes ; 16 chromsomes) Humans = 23 chromosomes (2 pairs of each --> 46 total) --> People thought that human would have a HUGE amount of genetic information (thought people would have >120,000 genes) 2003 - genome was complete and it was SHOCKING that humans only have 20,000 genes

Answer 10

Humans are more complex despite gene numver - WHY ; how does gene number correlate with complexity --> DON'T KNOW

Answer 11

Human genome = very complex Conatains: 1. Genes 2. Repetative sequences (LINES + SINES + LTRs + Transprosons + microsatilites) - Many things are reminents of retrociruses

Answer 12

Viruses in humans = drive evolution --> get expansion of genetic material + chnages in gene regulation + HGT See molecular artificats of evolutionary history by seeing reminenets of retrovirsues - Reminenets of retrovirsuses = LINES + SINEs

Answer 13

Chromosome number and chromosome arrangment = major driver of mammalial speciation Humans and chimps = similar BUT vary in chromosomes - If 99.9% identical then could make viable offspring (Ex. Neanderthols and sapeins repdoucing BUT this all breaks downonce chromosmoes change DNA can be the same BUT if the chromosomes look different = can't form viable offspring = have speciation - DNA in humans and chimps is almost idetofcal BUT chome have a chromsome split into twi = can't reproduce together

Answer 14

Gene density varies across species Image: - Shows Human genome Vs. Yeast - Green = coding exns ; Grey = introns ; Pink = LINE ; Orange = SINE - Can see that there is a lot of pink between inhidividual genes (have lots of LINE elements) Vs. Yeast have compact genes (genes are next to each other) - Genes are 500 BP apart - Easier to sequence yeast genome becasue there are less things between the genes (Compared to sequencing humans that has ALu and LINE elements between the genes)

Answer 15

Repetative DNA that populates the human genome - reminenet of ancient virus

Answer 16

Humans - 6 genes per megabase (mist DNA in humans is repeates (Mostly SINEs + LINES0 Fly - 80 genes per megabase Yeast - 549 genes per megabase

Answer 17

1. Tandem Reepeats 2. Interspersed repeat elements

Answer 18

A lot of human genome = tandom repeats Includes: 1. Salitile DNA repeates - centromeric DNA 2. Minisatelie DNA - Telemeric DNA 3. Microsatlite - Used for genetic fingerprinting ALL arise due to recombination or slipage during replication Often occur in cluster array Image - See seperare region on two chrosmomes that likley arose from replication mistake

Answer 19

Overall - selfish DNA elements interspersed throughout the genome - Arise due to transposition (artifcats of viruses) - propegate and spread throughout the genome Includes SINEs + LINEs - SINEs + LINEs = artifcats of viruses - Have different types of SINEs and LINEs (table - shows types and fraction in genome) - MOST SINEs = Alu family

Answer 20

Look like virsues - has multiple proteins (vrisues can make polycystronic trasncripts) - Virsues = can make polycystronic trasncripts --> virsues need to have compact genoomes to spread throughout popultion - reduce genome by putting open reaidng frames next to each other = virses have evoloved to have mutiple open reading frame exchnaged by frameshift event See artificacts of viruses in elements = shows they are viruses that littered the genome - Make up most of the genome ; most of genome is virus

Answer 21

Interspersed repeat elements (SINEs + LINEs) = major dirver of evolution --> selfish peices of DNA replicate and go to a new location --> make the genome bigger Interspersed repeat elements = Mechanism of speciation = changes in chromosome composition (expansion of chrosmome size or compaction of chromsome size) --> Interspersed repeat elements can chnage the chromsome = drive evlutiion - IF make the chromosome bigger = can't line up with predacessor = causes speciation - Viruses = responsible for a lot of evolution

Answer 22

Answer - 3 - Assumed the number of genes would correspond to the number of different proteins produced Assumed that because humans are complex (motile + have immune system) that we must make a TON of proteins to do everything

Answer 23

Gene number is a relativley poor correlate of organsim complexity (Complexity does NOT correlate with number of genes) Example (TABLE ON HIS SLIDES)- Plants = most number of protein coding genes C.elegans = has almost as many protein coding genes has humans Arabidopas = has more protein coding genes Corn = Has 100,000 protein coding genes (most)

Answer 24

Humans = do not have more protein coding genes BUT might still make more proteins Humans = 23,000 genes BUT can make up to 100,000 proteins BECAUSE we have split genes that are interupted by introns Can have alternative splicing: Interuption = creates a need for splicing --> exons do not have to be spliced togetehr in one way (can put they together in different ways) - Image - can have 2,3.4 or 1,2,5,6, = get two different proteins from 1 mRNA - 20,000 genes can make 100,000 proetins - creates alternativley spliced isoforms - 95% of intron containing protein coding genes undergo alternative splicing

Answer 25

Exons = encode polypeptide Introns = exoanses of non-coding DNA between exons Genes are translated as one unit (introns + exons) giving pre-mRNA --> pre-mRNA needs to be spliced and the exons need to be put together to translate polypetde

Answer 26

Exons tend to encode peptid units Example - exon might encode an alpha helix or a beta sheet (don't see alpha helix split between two exons) - Have modular unit of polypetide that gets put together with another modular unit

Answer 27

RNA recognition motif Have 2000 gene sthat bind RNA in some way (500 are common in a common binding motid called RRM) - Common RRM = all the the same --> didn't evove 500 times INSTEAD it evoloved and was copied in the genome THEN littel chnages occired to change specificty Exon sequences can be stitch together to create more diversity

Answer 28

Overall - Non-coding RNAs are an important compenent of the genome Have non-coding RNAs in ALL genomes (Humans + prokryotes + Archea) - Humans have lots of non-coding RNAs

Answer 29

Short non-coding RNAs = relativley well understood Includes: 1. rRNA - most abundent nucelic acid - Have a single gene locus for ribosomes that are reiterated 250X in genome (have 250 ribsome genes) 2. sn0RNAs - chrmical modification of rRNA - Small RNAs that direct enzymes to rRNA to modify them 3. snRNAs - splicing (bind to pre-mRNA to help with splicing) 4. siRNA + miRNA - RNAi - miRNA - small RNA that has translational control of mRNA 5. piRNAs - silencing of retrotransposons (keeps them from jumping around)

Answer 30

Long Non-coding RNAs are enigmatic - It is thought that even through there are deserts with no mRNA there are other types of RNA being made (including long-nc RNA) ~50,000 lncRNA trasncripts in humans Much lower abdince that protein coding transcripts (1-2 per cell) Some have well defined function (Ex. Xist)

Answer 31

Ribosoms = made in the nucleus BUT assumed in the nucleolus Nucleolus = section of nucelus where the ribsomes are made - All in an array in genome (all Eukaryotc genome) rRNA gene are in tandom array and all trasncribed - all transription and provess

Answer 32

Yeast - almost all nucelotodes are transcribed to mRNA or long nc-RNA (have trasncriotion of every moleculae)

Answer 33

Xist - dosage compensation In Humans have sexual dymorphism by the X chromosome - XX vs. XY - Have genetic imbalance ebtwen males have 1/2 the amount of X material --> imblance would be bad BUT this issue is soloved by expressing long-nc RNA in females (exoress Xist) Xist - randomly condenses chromatin in 1 female X chromsome (silences 1 X chrosome) = only 1 X chromosome is active = have balance - Reason all females are a mosaic of X chromosomes Every Eukaryote with sxual dymoprhism needs to deal with dosage compensation --> shows non-coding RNAs can be important in regulating the chromsome

Answer 34

Most metazoans solve impablance by silencing female X chromsome but insect dirve up the X chromsome 2 fold to balance out

Answer 35

8-15% of the human genome is under fucntional constraint

Answer 36

DNA is packaged Genomes can be rich in repetative DNA Genes can be discontinous Gene number is a poor correlate of biological complexity Number of genes does not indicate the number of proteins synthesized

Answer 37

1. By Developmental onset 2. Spatial expression patterns

Answer 38

Eukryotes = don't have operons but can organize DNA occruding to function Example - Hemaglobin has two types (alpha and beta types) - The two types are organized in the genome in a similar fashion so have embryonic and adult form - Chnage expressiion of hemoglobin depneding on age (baby is different than adults) BUT the genes are loctaed in the same area Proximity to the LCR proposed to guide the orger of globon gene expression during development

Answer 39

Genes can be organized accoding to spatial expression pattens - Genes can be arranged occurding to bpdy patterning - Order of genes on the chromosoms is the same as teh expresison of the genes in the developing embryo Have body patterning in ALL multi ceullar organisms - Genes involoved in bdy patterning locatiion = found on teh same chromsomes AND in a predictable pattern (true through lineages) Example - Drosophilla genes are similar to humans and mice (arranged in the same fashion) - Ancient way to arrange DNA

Answer 40

Regions of the nucelus are preferentially occupied by a particular chromosome - May affect regulation and might faciliatte their ability to match up in mitosis and meiosis Image - each chromosome is stained --> can see they go to different locations in the nucleus

Answer 41

TADs - Genomes are organized in Topoligically associated Domains (TADs)

Answer 42

Regions of the chromosome that bind to eachother (regions that are far away that bind to eachother?) - Allows for higher order regulation Found based on 3D structure of chromosomes To find them: Proteins bind to DNA in chromosomes --> cross link DNA --> cut DNA with rstcition enzymes --> NOW have two peices of DNA that are stuck because you criss linked them --> fill end and mark with biotin --> ligate --> purify and shear DNA ; pull now biotin --> sequence the two peices of DNA (see that they should not have been near each other) - Saw there is patterns to where chromosomes line up with themselves (called TADs) - Chromsome confirmation capture provides a contact map

Answer 43

TADs are conserved across all cell types and related species - Regions of TADs from diferent cells are the same - Humans and chimps have the same TADs

Answer 44

TADs = functional domains that have genes that are expressed similarly Rather than ranging genes linearly (like in operon) INSTEAD they are arranged in space by tethering them together Domains are close together because of chromatin (protein around DNA) - Protein brings peice of DNA close together -->THEN have transcription in envirnment of several genes of similar function

Answer 45

Genes can be organized on chromosome acoding to developmental pathway Each chromosome has its own teritpry A chromosome is made up of distict TADs TADs are deliminated by insulators that miantain their independece TADs are likley functional domains in the genome

Answer 46

TADs = regions of DNA that blocks transcription to other areas and gets localized transcriotion in that area

Answer 47

90s - used Sanger sequencing --> usses ddNTPs to get fragments of distcit length that you resolve on a gel Uses: 1. ddNTP - stops DNA replication 2. Primer 3. DNA polymerase Have 4 test tubes - each with a different ddNTP End get different strands of different sizes where you knwo the last nucleotode - Run segments ona gel = get ladder that is 1 nucleide different = can read sequuence IMAGE - shows 5' and 3 end AND dNTP which replaces 3' OH with H ; see that have different fragments where know the last nucleotide

Answer 48

Next gen = fancy sanger sequencing BUT next gen is not read by hand INSTEAd it ses computers Computing power allows us to sequence huge amounts of DNA in little bits = NGS

Answer 49

Random breakage by sonification THEN need to realign to reference genome 90s - sequencing by hand = someone needed to match everything to see what assmebled on what NOW - shotgun sequencing --> have sequencing fragments of DNA (few dozed nucleotides at a time) --> computer assmbles genome

Answer 50

NOW = very fast Took days-weeks to sequence COVOD (before would have taken 2-3 years to isolate and sequuence the virus) Because of sequencing = could develop he vaccine in 1 day (1 day to sequence clone into a vector) - Longest period during development = having the vaccine sit to make sure it is sterile

Answer 51

Detect RNA and DNA + modification of DNA/RNA + detect different types of RNA

Answer 52

Sample prep - Add dapdters to end of DNA fragemnts - Add motfies (indecies + reguon complemetry to oligions + one more) Cluerting - each fragemnt is amplified - - Hybriizetion ny one oligio - complementray to adpater readion on teh fragment - make dsDNA --> then denatured and teplate is washe --> amplify y bridge amiplifcation (binds to itehr oigio to make bring and then polymerase --> then denature --> now 2 copies of moclue) --> repeat process --> get amplification of the fragemmts --> reverse strands are cleaved and washed off Sequencing - extend the sequneicng primer --> floruencetley atged nucoetided ad t chain bease on template sequences - After each nucleotide comes = have excitation - flruencnt ddNTP -- have dsck pumoing in cnuetide sin regulated fashio (A --> G --> G --> T ) - camera at the top look sat flashed of light across al clusters - did billions of sewueincg read in minytes -- computer puts togteher nlines up and map against genome WATCH VIDEO NOTE - lecture stopped here BUT made flashcrads of rest of slides

Answer 53

Obtain DNA sequences from all genomes in a particular habitat

Answer 54

Sequencing reads used as a method of counting the numver of times a sequence appears

Answer 55

umans have 48 Mb of coding DNA (1.5% of genome_ Use oligionucletides as baits to hydridize to DNA of interest

Answer 56

To do - Prepare cDNA library and subject to NGS Sequence reads that corespond to segments of trasncripts in original RNA

Answer 57

Bisulfae converts C--> U (U is read of thymine)

Answer 58

Humans genome has 42% GC content Expected = 0.21 X 0.21 = 4.41% CpG BUT <1% is observed

Answer 59

Genome maps provide a framework for sequencing prohects There is NO such thing as THE genome sequence NGS allows sequencing of millions of molecules in parallel The human genome still has some sequence gaps CpG dinucleotdes are underrepresneted in the human genome

Answer 60

Sample prep 1. Add Adpatpters tp the end of teh DNGA fragments 2. Add the Sequence binidng site + Inidcies + regions complemenrray to the oligios Clutering - Isothermal amplification of the fragemnts 1. Region compleentary to the oligio binds 2. DNA plymerase extends and makes dsDNA 3. Unattached DNA strand gets washed away 4. Second end of DNA frament binds to 2nd oligo on slide (forms birdge) 5. DNA polymerase extends and makes dsDNA -> stands will seperate (both starnds are attached to the slide) 6. Repeat to keep amplifying 7. Reverse strand is washed away (only have the foward strand) Sequencing - 1. Extned the sequencing primer ; Nucleoodes have floruecent tages --> flruencent when added to the growing DNA strand - Base call is determined by teh wL and the sigla intesity - # of cylcles determiens the read length 2. Read is washed away 3. Index 1 primer binds (lower on starnd) --> extend primer --> index 1 read is washed away 4. Other end of opligio binds and forms bridge--> Index 2 primer binds 5. DNA polymerase extends and makes dsDNA 6. Foward strand is wahsed away 7. Complete the reverse read Anylysis - have foward and reverse reads that forms contnuogs that are assmbed onto a refernce genome - Samples are pooled based on unique indicies

Brainscape's Knowledge GenomeTM

Lecture #1 (Genome organization) Flashcards

Brainscape's Knowledge Genome^TM