BSC2010 EXAM 2 STUDY GUIDE
BSC2010 EXAM 2 STUDY GUIDE BSC 2010
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This 18 page Study Guide was uploaded by Tori Ruby on Saturday March 19, 2016. The Study Guide belongs to BSC 2010 at University of Florida taught by Staff in Winter 2016. Since its upload, it has received 228 views. For similar materials see Integrated Principles of Biology 1 in Biological Sciences at University of Florida.
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Date Created: 03/19/16
February 7, 2016 Reproduction: Problems to be solved o Transfer of Information Between parent and daughter cells Between parent and offspring (in multicellular organisms) o Apportionment of biological materials (e.g., organelles) Types of Reproduction o 1. Asexual (“clonal”) Reproduction Offspring (cell, individual) are genetically identical* to THE parent All prokaryotes**, many eukaryotes Occurs by binary fission in prokaryotes, mitosis in eukaryotes o 2. Sexual Reproduction TWO parents combine their genes in a stereotypical way (i.e., meiosis) Leads to recombination, i.e., offspring are genetically different from either parent ONLY occurs in eukaryotes Interphase (G1) o Each chromosome is a single, unreplicated double strand of DNA o One chromosome from each parent (Male, Female) forms a Homologous Pair (=“homologs”) o Chromosomes in nucleus, surrounded by nuclear membrane o Single centrosome S-Phase o After DNA replication, TWO “sister chromatids” are present for each homolog o Each sister chromatid is the SAME double-stranded DNA molecule o The two sister chromatids are attached by proteins o Centrosomes duplicate Early Prophase o Chromosomes condense o Mitotic spindle forms from centrosome o Centrosomes begin to migrate to poles of cell o Nucleoli disappear Mid-Prophase o Chromosomes fully condensed o Centrosomes complete migration to the poles o Nuclear envelope begins to degrade o Spindle fibers enter nuclear area from the pole o Kinetochores form at centromeres Prometaphase o Nuclear envelope completely degraded o Spindle fibers begin to attach at kinetochores o Sister chromatids attached to opposite poles Metaphase o Spindle fibers attached to centromere at kinetochore o All sister chromatids attached to opposite poles o Chromosomes migrate to center plane of cell, “metaphase plate” Anaphase o Protein bond between sister chromatids degrades o Sister chromatids separate, migrate toward opposite poles o Poles move farther apart as non-kinetochore spindle fibers lengthen Telophase o Non-kinetochore spindle fibers continue to elongate cell o Nuclear envelopes begin to form at poles o Chromosomes de-condense back into chromatin o Nucleoli re-form, cytokinesis begins Eukaryotic Cell Cycle Control o Cell replication must be under precise control. o If unicellular organisms had no control over reproduction, they would exhaust their resources and starve to death. o Signals from external environment o In multicellular organisms, cell reproduction must be controlled for proper development. o Internal signals, e.g., growth factors February 11, 2016 Cell control o 1. CDKs act on cell-cycle regulators o 2. Example: G1-S checkpoint Sexual Life Cycles – Human Example o 46 Chromosomes o 22 Homologous Pairs of autosomes Same length • Same centromere position • Same sequence (+/-) • Same set of genes o • One pair of sex chromosomes • Females XX, Males XY • Only small region of sequence homology Meiosis o • RECALL: Function of MITOSIS is to faithfully replicate the parental genome in each daughter cell with no change in information content o • Proximate function of MEIOSIS is to produce haploid cells from diploid cells o • Ultimate function of MEIOSIS is to generate genetic variation upon which natural selection can act Sexual Life Cycles – Diplontic (e.g., animals) o Free-living stage is diploid o Gametes formed by meiosis o Haploid gametes merge genomes to form diploid zygote (“syngamy”) Sexual Life Cycles – Alternation of Generations (e.g., Plants) o Diploid sporophyte forms haploid spores by meiosis o Spores form gametophyte by mitosis o Gametophyte forms gametes by mitosis o Gametes merge to form diploid zygote Sexual Life Cycles – Haplontic (e.g., Fungi) o Free-living, multicellular organism is haploid o Gametes formed by mitosis o Gametes merge to form diploid zygote o Zygote undergoes meiosis to form haploid cells Meiosis – an overview o Interphase 1: G1 phase – • Begin with two homologous chromosomes (out of n homologous pairs) • DNA content = 2C • Ploidy = 2n (diploid) o Interphase 1: S-phase • Chromosomes replicate • DNA content = 4C • Ploidy = 2n o “Meiosis I” • Homologous chromosomes separate • Cell Division* #1 • Result is TWO haploid (ploidy = n) cells with TWO SISTER CHROMATIDS of ONE of the two homologs o “Meiosis II” • Sister chromatids separate • Cell Division # 2 • Result is FOUR haploid daughter cells, each with a single unreplicated chromosome (= 1C) o • Each cell contains ONE member of each homologous pair of chromosomes Meiosis I – early Prophase I o Homologous chromosomes pair o Synaptonemal complex (proteins) attaches homologs • “synapsis” o Homologs form tetrad Meiosis I – late Prophase I o Chromosomes cross over, form “chiasmata” o Exchange of DNA between homologs occurs at chiasma o Spindles form and attach to kinetochores as in mitosis Meiosis I – Metaphase I o Chromosomes lined up on metaphase plate in homologous pairs o Spindles from one pole attach to one chromosome of each pair o Spindles from the other pole attach to the other chromosome of the pair Meiosis I – Anaphase I o Homologous chromosomes separate and move along spindle fibers toward pole o Sister chromatids remain attached at centromeres o Note that recombination has occurred Meiosis I – Telophase and cytokinesis o Homologous chromosomes reach (opposite) poles o Each pole has complete haploid complement of chromosomes o Each chromosome consists of two sister chromatids Meiosis II – Prophase II o Spindle forms o Chromosomes move toward metaphase plate Meiosis II – Metaphase II o Chromosomes reach metaphase plate, as in mitosis o Kinetochores of sister chromatids attach to spindle fibers from opposite poles Meiosis II – Anaphase II o Centromeres of sister chromatids separate o Sister chromatids move toward opposite poles Meiosis II – Telophase and cytokinesis o Mechanism as before o Note that now FOUR HAPLOID DAUGHTER CELLS formed from each parent cell o Note that some chromosomes are recombinant, some are no Apoptosis - Programmed Cell Death o At certain points in development, cells are programmed to die o Failure of apoptosis leads to developmental abnormalities (e.g., webbed hands and feet in humans) o Cells with irreparable DNA damage (can lead to cancer, abnormal development) also subjected to apoptosis o "Suicide" proteins (caspases) are always present in inactive form Regulation is post-translational, not of transcription February 17, 2016 Dihybrid cross- dependent assortment o Predict ¾ round and yellow, ¼ wrinkled and green o Not what Mendel observed Independent assortment o Four combos of alleles in gametes o All equally likely o 9:3:3:1 ratio expected o This is what Mendel observed Mendel’s two laws o 1. Law of segregation If the locus is heterozygous, half of the gametes get one allele, half get the other allele o 2. Law of independent assortment- multiple loci Alleles at each locus segregate independently of one another Probability theory: the chance of something happening, relative to all possible outcomes o Independent: outcome of one event has no effect on the outcome of the other Ex. Rolls of a die, flips of a coin Probability of both events Pr(A and B)= Pr(A)x Pr(B) Probablility of either or two events happening Pr(A or B) = Pr(A)+ Pr(B) Incomplete dominance o Ex. Snapdragons o Two alleles: C^R and C^W o Heterozygotes are intermediate between two phenotypes Co-dominance o One locus, two alleles o Ex. M/N blood groups o M/N locus encodes glycoprotein antigen that occurs on the surface of the human red blood cells o Heterozygotes show both phenotypes Multiple alleles o Most genes have more than 2 alleles o ABO blood type One gene, 3 alleles: I^A, I^B, i (i is null) I^A and I^B are co-dominant, i is recessive Pleitrophy: one gene effects multiple characters o Ex. Tyrosinase (Tyr-albinism) o Ex. Sry (sex- determining locus in mammals) Epistasis: an allele at one locus affects the phenotypic effects of an allele at the other locus o Ex. Mouse coat color One locus controls hair pigment color Black, brown; black is dominant (BB, Bb, bb) Second locus controls pigment deposition Pigment dominant to no pigment (CC, Cc, cc) Dihybrid cross (BbCc x BbCc) Polygenic inheritance o Most traits do not have a simple genetic basis o Called “quantitative traits” or “complex traits” o Often continuous traits Height, weight, skin color Ex. Many chronic diseases o Consider the offspring of a mating between two individuals heterozygous at all loci- AaBbCc x AaBbCc What fraction of gametes will be ABC? 1/8 What fraction of gametes will be AABBCC? 1/64 Environment effects on phenotypes o Many traits not completely determined by genes Height, weight, skin color Many chronic human diseases, ex. Coronary heart diseases o Ex. Light intensity on plant height o Called “genotypes by environment interaction” Human genetics- pedigree analysis o Can’t do controlled crosses with humans o We track phenotypes on “pedigree” Genetics of disease o Recessive disorders Most diseases are recessive Homozygous recessive have disease Heterozygous are carriers March 7, 2016 Mutation: change in the composition of the genome from parent to daughter strand o (total genomic damage-repaired damage) = mutation o Occur in both somatic and germline cells (only considering heritable mutations- which occur in germline cells) o Is the ultimate source of genetic variation (!) Categories of mutations o Nucleotide sequence mutations o Chromosomal rearrangements o Transposable elements Types of nucleotide sequence mutations o Point mutation (+ base substitution) o Insertions/deletions (“indels”) o Duplications Point mutations o Substitution of one nucleotide for another at a homologous site o Rates vary- generally around 10^-8 to 10^-10 per site per generation, varies by taxon o Occur anywhere in the DNA sequence Rate may (and does) vary with genomic text Chemical types of point mutations o o Transitions are more common- magnitude depends on gene and taxon The genetic code o Mutation- “silent” or “synonymous” substitution o Substitute a purine for pyrimidine or pyrimidine for purine Mutation- “Replacement” substitution o Substitute purine for purine or pyrimidine for pyrimidine Mutation- insertion/deletion (“indels”) o Deletion: remove a pyrimidine or purine o Insertion: dd a pyrimidine or purine o Can be one or more nucleotides Mutations occur because o Spontaneous errors in DNA replication (mismatches) o Mutagenic damage to DNA (replicating or non-replicating) o “Selfish” elements replicate themselves, often to the detriment of the host genome Causes of mismatches o Polymerase base misincorporation DNA polymerases have 3’->5’ and/or 5’->3’ exonuclease (proofreading) function o Tautomerization o Spontaneous deamination (e.g., deaminated cytosine is uracil, leads to C/G-> U/A -> T/A DNA repair o First line of defense: DNA polymerase DNA polymerase III has a “proofreading” capability If the wrong nucleotide is entered, DNA synthesis stops, the offending nucleotide is removed, and synthesis resumes Mechanisms of mutagenic damage o Base analogs: purines/pyrimidines that mimic legitimate bases pair differently o Direct damage to DNA: e.g. Deamination, alkylation, intercalating agents, UV o Indirect damage to DNA: e.g. Agents that generate oxygen free radicals Oxygen free radicals predominantly cause G/C->A/T via 5- hydroxyuracil from cytosine deamination and G/C-> T/A via 8- oxoguanine Mechanisms of DNA repair o Direct repair: damage repaired directly o Excision repair: damaged DNA excised and repaired using undamaged strand as template; base excision, nucleotide excision o Mismatch repair: recognizes misincorporated bases, removes the mispaired base, and repairs using the parent strand as a template o Recombination repair: (double stranded break repair): employs the recominational machinery o Transcription- coupled repair: (ER, specific to transcribed strand during transcription) Insertions/Deletions (indels) o Insertions Addition of nucleotides to a sequence o Deletions Deletion of nucleotides from a sequence o Result from “replication slippage” when replicating DNA reanneals at short tandem repeats Non-homologous (ectopic) recombination Chromosomal aberrations and their consequences o Non-disjunction Homologous chromosomes do not separate at meiosis I Sister chromatids do not separate at meiosis II Consequences If fertilization with a gamete with abnormal ploidy occurs, result is offspring with abnormal chromosome number, called “aneuploidy” Polyploidy o Sometimes organisms get an entire extra (haploid) set of chromosomes o 3 sets are called “triploidy” Can occur by fertilization of a diploid egg in which all chromosomes underwent non-disjunction o 4 sets are called “tetraploidy” E.g. Zygote fails to divide after DNA replication; mitosis leads to a 4n organism Duplications o Result from non-homologous recombination o Probably very important for evolution of new genes Chromosomal rearrangements o Typically from non-homologous recombination or double-stranded breaks in the chromosome o 2 types Inversions Once inversion occurs, recombination is usually prevented because recombinant gametes cannot get a full complement of genes Translocations Pieces of non-homologous chromosomes break and fuse together Often cause improper segregation of chromosomes at meiosis Transposable elements: “jumping genes” o Transposable elements are genetic elements that contain the information for their own replication o Insert into new locations in the genome Conservative transposition (cut and paste) Replicative transposition (copy and paste) o Unlike episomes or viruses, TEs never exist independent of the host genome o Most recombination requires complementary base-pairing of homologous (sequence-similar) regions of DNA Meiosis (in eukaryotes) Transformation Generalized transduction Conjugation o Transposition is a form of non-homologous recombination o TEs can move to novel sites in the genome o The simplest TEs consist of A gene that encodes an enzyme for excision an insertion of the TE sequence; transposase in DNA TEs A recognition sequence that the enzyme recognizes as the boundary of the TE o In prokaryotes Insertion sequences A transposase gene A recognition sequence that is an inverted repeat Transposase recognizes boundaries of TE inverted repeats and cuts the TE out of the donor site Transposase cleaves chromosome at target side via endonuclease Transposon is joined to the single-stranded ends at the target site Gaps are filled in by DNA polymerase I and DNA ligase Result is a new copy (copy and paste) or moved copy (cut and paste) of the TE o Composite transposable elements Contain additional genetic material besides transposase, IRs Structure consistent with two ISs close together in genome One transposase may not be functional March 9, 2016 Transposable elements (TEs) o DNA transposons Composite transposable elements Contains additional genetic material besides transposase, IRs Structure consistent with two ISs close together in genome One transposase may not be functional Prokaryotic gene regulation: metabolic control o Metabolism can be defined as the total of the chemical reactions of a cell (or organism) Must be controlled, or chaos ensues o Anabolic metabolism: building large molecules out of small molecules; requires energy input o Catabolic metabolism: breaking down large molecules into small molecules; releases energy o Direction of flux through metabolic pathway depends on cellular needs o Flux through a metabolic pathway can be regulated in two ways Activity of the enzymes themselves Regulation of enzyme activity o Competitive inhibition: inhibitor binds to active site o Non-competitive inhibition: inhibitor binds to some other part of the cell o Allosteric regulation: multiple subunits o Feedback inhibition Metabolic pathway switched off by the end- product o Cooperativity Active form stabilized by substrate Expression of genes that encode enzymes Regulation of gene expression: operon o In prokaryotes, functionally related genes often are clustered together in the genome, called “operons” o Under the control of a single promoter sequence, so are transcribed together o Single “switch” sequence controls transcription of the whole operon; called an operator o Operator controls access of RNA Pol to genes o Repressor proteins controls access of Pol to operator o Repressor protein encoded by a “regulatory” gene o Presence or absence of an end-product or precursor controls the operon “on-off switch” o Example: the E. coli trp operon o Trp operon is repressible because transcription is repressed by the presence of an end-product Repressible operons usually function in anabolic metabolism o An inducible operon becomes transcriptionally active by the presence of a precursor Example: E. coli lac operon Negative v. positive o Trp and lac operons are negative regulation: transcription switched off by active repressor o Positive regulation: transcription switched on by activator molecule Operon control: repressible v. inducible o Repressible operon: end-product turns the operon off o Inducible operon: substrate turns the operon on March 11, 2016 Eukaryotic genomes: physical structure o Eukaryotes face different organizational challenges tan prokaryotes Cells are much larger and more complex Many eukaryotes are multicellular; cells have to differentiate Genomes are larger and more complex o Chromatin: the complex of DNA and proteins that together constitute the chromosome o Problem: how to pack an enormous amount of DNA into a cell o Chromatin is sequentially packaged o Level 1: Chromatin fibers (10 nm) Histones are positively charged proteins that bind to chromosomal DNA Remain complexed with DNA throughout the cell cycle Changes in shape and orientation of nucleosomes influence accessibility of DNA to RNA polymerase o Level 2: 30 nm fiber Histone tails interact with linker DNA to form loops in the 10 nm fiber o Further folding of the chromatin results in mitotic condensation of chromosomes o Heterochromatin remains densely coiled during interphase o Euchromatin uncoils during interphase Control of gene expression o All cells in a multicellular organism contain the exact same genetic compliment o Any cell expresses only a small fraction of its genes at any given time o Regulation of the expression of specific genes most common at the level of transcription o Gene expression typically depends on the cell receiving some signal o Upon receipt of a signal, regulation can potentially occur at many steps o 1. Regulation of chromatin structure- modification of histones Acetylated histones: -COCH3 groups attached to lysine in histone tails Acetylated histones cannot bind nucleosomes, chromatin de- condenses Conversely, methylation of histone tails causes chromatin to condense Each histone tail contains several lysine residues that can be acetylated or methylated Information stored in pattern of acetylation/methylation o 2. DNA methylation DNA of most higher eukaryotes is methylated in place DNA methyltransferases, other demethylating enzymes Most common is methylated cytosine Methylation usually represses expression Can be preserved across replication; methylation enzymes methylate daughter strand Information contained in methylation can be passed to offspring; “epigenetic” inheritance Genomic imprinting Basic assumption of Mendelian genetics is that the effects of an allele do not depend on whether the allele was inherited from Mom or Dad Sometimes, effects of an allele depend on the parent from which the allele was inherited Imprint is often due to methylation of one allele Inherited imprint is removed in gamete producing cells Now, sex specific imprint is applied in the germ cells Gametes have appropriate imprint Dosage compensation of sex-linked genes In organisms with chromosomal sex determination, the homogametic sex typically has two copies of sex-linked genes and the heterogametic sex only has one copy This can cause a problem, because all else equal, the homogametic sex will have twice as much gene product expressed as the heterogametic sex Dosage compensation has evolved to equalize gene expression between the sexes In placental mammals, one X chromosome in somatic cells in females is inactivated, X inactivation Mechanism of X inactivation is expression of a functional RNA gene called Xist Xist RNA coats the X to be inactivated, causing the whole X to be heterochromatin The targeted X is random o 3. Regulation of initiation of transcription Transcription factors associated w/ RNA Pol are necessary for transcription of all genes The general transcription machinery usually transcribes genes at a low rate Efficient transcription requires specific transcription factors The same gene may be controlled by different enhancers in different cell types, developmental stages, etc Some transcription factors act as activators Activators recruit proteins that acetylate nearby histones Other TFs act as repressors Repressors recruit proteins that deacetylate histones 1. Activator protein binds to enhancer 2. Binding of activator causes DNA bending, activators brought in proximity to promoter 3. Activators bind to other TFs, allow formation of an active transcription complex on the promoter Coordinated control of gene expression o In general, many genes need to be co-regulated Eukaryotes typically do not have operons Relatively few sequences in control elements Enhancers typically composed of about 10 control elements Specificity of control elements depends on the particular combination rather than a single unique sequence o Co-regulation usually depends on association of a specific set of control elements with each gene in the co-regulated group Post transcriptional processing o Gene expression ultimately depends on the amount of gene product o Gene expression can be regulated post-transcriptionally by Alternative splicing mRNA degradation microRNA, small interfering RNA Regulating translation “Maternal effect” genes lack a poly-A tail Post-translational control Why introns o Allow a single gene to encode >1 polypeptide March 14, 2016 Viruses o Viruses are obligate parasites of cellular organisms o Consist of nucleic acid genome (DNA or RNA) enclosed in protein o Use the host’s cellular machinery to replicate o Viral genome encodes the information necessary to replicate Phage reproduction: lytic and lysogenic o Bacteriophage are viruses that attack bacteria o Lytic reproduction kills host by lysis Phage that reproduce only by lytic reproduction are called virulent 1. Phage binds to host’s surface receptor 2. Phage injects DNA into the cell 3. Host’s DNA is hydrolyzed (degraded) 4. Host’s metabolic machinery used to make phage protein and DNA from free nucleotides and amino acids 5. Phage encodes lysozyme production, cell lyses, new phage released o Lysogenic reproduction replicates phage genome along with host genome without killing host Phage that can reproduce by lytic or lysogenic reproduction are called temperate 1. Phage attaches, injects DNA 2. Phage DNA circularizes 3. Phage DNA integrates by crossing over via phage integrase. Integrated phage called prophage: lives for a long time 4. Occasionally, prophage is removed from host genome via phage excisionase; lytic cycle ensues Genetics of prokaryotes o Bacteria have a single circular chromosome o Many bacteria have plasmids: self-replicating elements that can integrate into the chromosome and which contain only a few genes o Bacterial genomes contain between 500 and several thousand genes o Prokaryotic genome is much smaller than eukaryotic genomes o Very few introns; no splicosomes Prokaryotic recombination o Bacteria reproduce by clonal fission (asexually) o Yet, recombination is common Transformation Uptake of naked DNA Transduction Phage mediated DNA transfer Conjugation “mating” between bacteria Inference of recombination o 2 strains of E. coli; arg- and trp- o Plate on minimal medium o Neither strain grows by itself, but a mixture grows o Must be recombination not mutation because the mutation rate is known Recombination o Transformation Takes up foreign DNA double stranded RNA Integrated into bacterial genome at a site of sequence homology Many species of bacteria have receptor proteins on the cell surface that function in DNA uptake Uses homologous sequence to replace a strand of host DNA One daughter has the donor strand, one has the host strand o Phage-mediated exchange 1. Phage infects bacteria 2. DNA hydrolyzed, phage DNA replicated 3. Fragment of bacterial DNA incorporated into phage 4. Phage infects new host 5. Crossing over can occur between bacterial DNA fragment and homologous region of chromosome 6. Recombinant genotype results o Recall lysogenic phage Prophage exits host genome before entry into the lytic cycle Occasionally the prophage takes some of the host genome with it Results as before, recombination can occur Comparative genomes: sequence organization o Prokaryotes most of the genome encodes either protein, functional RNA, or regulatory sequence o In multicellular eukaryotes, the vast majority of DNA does not code on functional gene product o Much non-coding DNA is not “junk DNA” (but some may be) o Rate of increase of amount of DNA from prokaryotes to single- celled eukaryotes to multicellular eukaryotes is much greater than the increase in number of gametes TE related sequences o Many TE related sequences are TEs that have lost the ability to transpose Called non- autonomous elements Can sometimes be mobilized by enzymatic machinery of “live” TEs Other repetitive DNA o ≈15% of human genome is repetitive DNA not related to TEs o Mostly due to errors in DNA replication and/or recombination Large-segment duplications Multi-gene families o Most eukaryotic genes present in only one copy o Some genes present in >1 copy (eg. rRNA) o Some genes closely resemble each other in sequence Genes that closely resemble each other probably resulted from an ancestral gene duplication May be clustered or dispersed in genome Fuctional genomics: quantitative trait locus (QTL) mapping o Hypothetical example: there is a genetic variation among inbreeding lines of mice in the susceptibility to infection by staphylocus aureus o Question: how to find the genes that confer susceptibility? o Cross a susceptible line to a resistant line o F1s will be heterozygous at all loci o F2s will segregate different combinations of alleles o Genotype the lines at many marker loci Functional genomics: genome- wide association study (GWAS) o Example: susceptibility of schizophrenia in humans o Schizophrenia seems to happen in families more often than expected by chance, but it’s obviously not due to one or a few Mendelian loci o How to track down genetic causes? You can’t do a controlled breeding with humans o Solution: Let nature provide the meiosis for you! o Genotype a large number of individuals at a large number of marker loci, “single nucleotide polymorphisms” (SNPs) March 16, 2016 Purposes of biotechnology o 1. Establish the genetic basis of phenotype Eg. Find mutations that cause colon cancer o 2. Manipulate the genetic basis of phenotype Eg. Fix mutations that cause colon cancer Methods o Restriction enzymes/gene cloning Restriction enzymes allow for the manipulation of DNA in a repeatable predictable fashion Cloning vector – DNA or RNA molecule (e.g., bacterial plasmid or virus) capable of independent replication Plasmid – Small, double-stranded ring of DNA, found in the cytoplasm of bacteria and capable of independent replication Basic strategy is to ligate (link) target DNA to vector DNA, so that when the vector replicates, so does the linked target DNA o Polymerase chain reaction (PCR) Isolate and amplify a specific DNA sequence o Transformation/transfection/transduction o Expression vectors o Genome editing with CRISPR/Cas9
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