Topic 9: Genetic Evolution and the Study of Genes Flashcards

Question

Neutral mutations

Answer 1

* do not affect the cell * in unimportant regions of the genome, silent mutations, difference in amino acid (missense) doesn't alter the protein function

Answer 2

Studying comparisons between genomes (when there's a change, shows when different organisms genomes diverged)

Answer 3

* homologous genes: similar nucleotide sequence because of common ancestry (can see how many differences arised over time)

Answer 4

Codes for an essential protein or RNA molecule * mutations are typically disastrous

Answer 5

Genes that code for ribosomal RNA -> all cells need, if rRNA not made properly, a cell will not survive

Answer 6

* diagram depicting the evolutionary relationships among a group of organisms * rRNA genes are so highly conserved we can make trees relating all domains of life (even distantly related organisms) -because if rRNA changed the organism wouldn't function

Answer 7

How many changes occured over time

Answer 8

Regions in which corresponding genes are strung together in the same order in multiple species

Answer 9

* Elimination of individuals carrying mutations that interfere with important functions (things that are the same between organisms, they are important * mutations in regions of the genome that do not tolerate mutation

Answer 10

* less than 1% is protein-coding exons -20 000 genes -5000 genes for functional RNAs * 4.5% of the genome is highly conserved (across all humans bc humans with changes would've died) * 5% of the genome has reduced variation between humans * therefore, only about 10% of the genome contains sequences that truly matter for survival (in the other 90% doesn't matter if we get mutations - this is where variation comes from) * differences between individuals are due to single nucleotide polymorphisms (SNPs)

Answer 11

single nucleotide polymorphisms (SNPs)

Answer 12

* cut DNA into fragments at specific nucleotide sequences (bc the chromosome is too big to study all at once so we must cut it up) -specifically restriction nucleases (cut up nucleotides) -4-8 nucleotide pairs -made naturally by bacteria -each enzyme is specific to the sequence it cuts (we choose one that cuts where we want) -can give straight cuts or staggered cuts creating sticky ends -more often we want staggered ends

Answer 13

* DNA fragements separated based on length * put in agarose or polyacrylamide gels * DNA moves from the negative electrode to the positive electrode (bc DNA has overall negative charge, it repels negative electrode) * DNA is labelled or stained -stained with ethidium bromide (most commonly) to fluoresce under UV light * you can then isolate a DNA fragment by excising the desired band with a scalpel

Answer 14

* inserting fragments into a vector (carrier)

Answer 15

* piece of DNA that can be copied inside cells (carry the gene of interest)

Answer 16

circular piece of DNA that contains its own replication origin (can cut with same restiriction enzyme we cut our DNA with, so plasmid and our gene have same sticky ends to complementary bp)

Answer 17

Joins together the pieces of DNA

Answer 18

* ability of some bacterial cells to take up DNA molecules from their surroundings (form of horizontal gene transfer) * DNA fragment of interest is produced in large amounts in individual cells * Cell are lysed and recombinant plasmids are collected

Answer 19

Collection of plasmids inside bacterial cells * represents the full genome of the organism (genome cut up and put in plasmids)

Answer 20

Contains only the protein-coding sequence of genes * cDNA = Complementary DNA * DNA is copied from the mRNAs present in a cell -use mRNA as a template to make a ds DNA copy with reverse transcriptase and DNA polymerase - which can then be put in plasmid) * DNA we're making lacks introns and other noncoding sequences (made from final mRNA, not pre-mRNA -better bc bacteria can't cut out introns)

Answer 21

* Billions of copies of a nucleotide sequence can be generated in a few hours * sensitive: detects and amplifies trace amounts of DNA * there is selectivity of DNA hybridization

Answer 22

* hybridization: ability of nucleic acid strands to bind to each other due to complementary base pairing * we construct primers specific to the gene we want to amplify - DNA primers are used to attach to the genes of interest in the DNA or RNA sample

Answer 23

1. Heat to separate strands (breaks the H bonds that hold the strands together) 2. cool it back down to allow primers to stick on to ss DNA 3. DNA synthesis: primers provide scaffold for DNA polymerase to build new strand -have 2 ds DNA now (DNA polymerase used is taq polymerase comes from archaea that lives in hot conditions so it can function when heated in first step)

Answer 24

* detection of infectious particles: using primers targetting genes specific to the suspected pathogen -track epidemics (can amplify virus in a swab sample amplify it and compare it to noninfected sample to determine if affected or compare it to different strains to determine strain, can also find where people are more affected by infection by checking waste waters and amplifying it (virus would be in waste of affected person) -detect bioterrorist attacks (can tell where it's coming from -like anthrax sent in envelopes) -test food products: if there's contaminants can determine what type of contaminants * forensics: -isolate DNA from small traces of blood or tissue -DNA fingerprint: unique human DNA sequences (differ from one human to another), use primers that target genes that are highly varied in the human pop'n

Answer 25

* type of DNA sequencing * dideoxyribonucleoside triphosphates (2 dna nucleoside) -chain terminating nucleotides (lack 3' OH -where another nucleotide would be added) -each terminator nucleotide labelled with a different fluorescent dye * DNA polymerase adds regular nucleoside triphosphates and terminator ones and its just by chance which is added * generates a collection of DNA fragments terminated at different positions * detector records the fluorescence to determine a nucleotide sequence

Answer 26

if you're only sequencing 1 gene, its cheaper

Answer 27

* DNA libraries made by PCR amplification of DNA fragments bound to solid supports (glass slides or beads) * when we go through amplification: -copies stay close to the parent molecule -so clusters are all sequenced at the same time

Answer 28

Illumina sequencing (most common way) * adds removable fluorescently-labelled chain-terminating nucleotides * after picture is recorded (of fluorescence), the label and chain terminator are stripped away, and it keeps going on the same strand Single-molecule real-time sequencing (SMRT) (not as common) * DNA polymerase, DNA template, primer anchored together with dNTPs * attachment of each nucleotide is determined one at a time, reads each nucleotide as it gets added * smarter bc doesn't need to add chain terminating nucleotides first

Answer 29

When you want to sequence an entire genome (newer method)

Answer 30

determines which genes are being expressed by analyzing which RNAs are being produced (bc if RNA produced -> gene was transcribed) -makes cDNA library using reverse transcriptase on RNA to make DNA -sequencing of cDNAs provides a quantitive analysis of the transcriptome

Answer 31

* complete set of RNAs produced by a cell under a certain set of conditions (if we know RNA is being transcribed in a certain cell, we can do a protein analysis to figure out which transcription regulators are in high vs low amounts)

Answer 32

Nucleic acid sequences are visualized in their normal location * single strand DNA and RNA probes labelled with fluorescent dyes or radioactive isotopes detect complementary nucleic acid sequences in a cell or tissue * allows us to see how transcription regulators guide development

Answer 33

* tells us where translational control is if there's RNA sitting there with translation turned off), and tells us which mRNAs are being translated * cell or tissue extracts are exposed to ribonucleases (break down mRNA) * mRNA sequences coated in ribosomes are not digested (the ones being translated bc ribosomes block so ribonucleases can't get past) * cDNAs of RNA that was being translated are studied (after ribosomes are stripped off)

Answer 34

* Gene of interest is replaced with a protein that can be easily monitored * used when we have a hard time seeing gene * Green fluorescent protein (GFP) -most common reporter gene used * study regulatory sequences that control the gene's expression (can use methods to turn regulatory sequence on or off, allow for activator to bind or not, put TF in or don't: and see if GFP is expressed, if it is it'll glow -so we can see how different things affect expression

Answer 35

what happens to an organism when a gene is inactivated (if we induce a mutation wat affect does it have on cell)

Answer 36

RNAi: method for silencing genes

Answer 37

* introduce a ds RNA molecule that matches the gene of interest * when introduced (RNA interference kicks in (bc it recognizes itself didn't make it) * the cell constructs siRNAs which hybridize with the target DNA sequence -this covers up the target DNA sequence so we are silencing that gene, stopping it from being transcribed -> we can see what affect not having access to a gene has -like if we think a certain disease is caused by a certain gene malfunctioning we can cover it up and see what it does -or if it's expressed all the time when it shouldn't be, if we silence it, we can see if the problem will be gone

Answer 38

altered gene is inserted into the genome of reproductive cells (egg, sperm or germline cells that make egg sperm) so it can be inherited (so we get a whole lineage that all have same gene altered to study affect)

Answer 39

activities of a gene can be eliminated entirely -not replaced, taken -lets us see how important gene really is, if organism can't survive without it = very important

Answer 40

* Cas9 enzyme produces a double-strand break in DNA -not sequence specific like other restriction enzymes -provide Cas9 with a guide RNA molecule (like telomerase has) to bind the DNA at a complementary sequence * homologous recombination kicks in to try and repair ds break: altered version of a gene serves as a template for homologous repair of the gene we have cut with Cas9 -replaces the normal gene with the altered gene

Answer 41

* have catalytically inactive Cas9 (not able to cut) fused with transcription activator and guide RNA to guide it to the specific sequence to see if that certain activator turns on gene * catalytically inactive Cas9 fused with transcription repressor to see if that repressor turns off gene

Answer 42

* treat disease: replace malfunctioning gene with a functional gene -could eliminate disease, not used yet but hopefully in future * use of model organisms to study mutated genes -if we replace mutated gene with functional gene, does that fix the disease -or insert mutated gene into model organisms to work on treatments for disease, or meds: this is alr being used * improvement of agricultural crops -genetically modified: alr being used -change nutritional concentrations -Resistance to pests, viruses, extreme weather -produce nutrients the crop did not normally make * produce any protein in high amounts for use in clinical applications and structural and biochemical studies; alr being used

Topic 9: Genetic Evolution and the Study of Genes Flashcards

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