The Molecular Basis of Inheritance Flashcards

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1
Q

scientists believed that proteins made up genes/inherited material

A
  1. proteins are major component of all cells
  2. complex macromolecules in seemingly limitless variety + great specificity of function
  3. a lot known about structure, very little known about DNA
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2
Q

Griffith (1927)

A
  • experiments with different strains of bacterium
  • some strains virulent and causes pneumonia, some strains harmless
  • discovered that bacteria have ability to transform harmless cells into virulent ones by transferring some genetic factor from one bacteria cell to the other
  • bacetrial transfromation + transformation experiment
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3
Q

Avery, MacLeod, McCarty (1944)

A
  • published findings that Griffith’s transformation factor is DNA
  • DNA was agent that carried genetic characteristics
  • provided direct experimental evidence that DNA, not protein, was the genetic material
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4
Q

Hershy and Chase (1952)

A
  • experiments that had strong support to DNA theory
  • tagged bacteriophages with radioactive isotopes (32P and 35S)
  • proteins contain sulfur but not phosphorus (DNA vice versa) so 32P labeled DNA of phage viruses while 35S labeled protein coat of phage viruses
  • when bacteria infected with phage viruses, 32P always entered bacterium while 35S remained outside
  • proved that DNA from viral nucleus, not protein from viral coat, was infecting bacteria and producing thousands of progeny
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5
Q

Rosalind Franklin (1950-53)

A

carried out X-ray crystallography analysis of DNA that showed helix

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6
Q

Watson and Crick (1953)

A
  • **proposed double helix structure of DNA **
  • used data from other scientists to build their model
  • 2 major pieces of data used were the biochemical analysis of DNA (Erwin Chargaff) and X-ray diffraction analysis of DNA (Rosalind Franklin)
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7
Q

Meselson and Stahl (1958)

A
  • proved that DNA replicates in semiconservative fashion (as Francis Crick predicted)
  • cultured bacteria in medium with heavy nitrogen, allowing bacteria to incorporate heavy nitrogen into DNA as they replicated/divided
  • bacteria then transferred to medium with light nitrogen and replicate/divided once
  • bacteria then spun in centrifuge and found to be midway in density between bacteria grown in heavy/light nitrogen
  • new bacteria contained DNA consisting of 1 heavy strand and 1 light strand
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8
Q

Structure of DNA

A

-double helix like twisted ladder, consist of 2 strands running in opposite direction (5’ to 3’ and 3’ to 5’)
- polymer consisting of repeating units of nucleotides, these consist of** 5-carbon sugar (deoxyribose), a phosphate, and a nitrogen base**
- carbon atoms in deoxyribose numbered 1 to 5
- four nitrogenous bases (adenine, thymine, cytosine, guanine: adenine and quanine purines, thymine and cytosine pryimidines)
- nitrogenous bases paired by hydrogen bonds: adenine bonds by double hydrogen bond to thymine, cytosine bonds by triple hydrgoen bond to guanine

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8
Q

Structure of DNA

A

-double helix like twisted ladder, consist of 2 strands running in opposite direction (5’ to 3’ and 3’ to 5’)
- polymer consisting of repeating units of nucleotides, these consist of** 5-carbon sugar (deoxyribose), a phosphate, and a nitrogen base**
- carbon atoms in deoxyribose numbered 1 to 5
- four nitrogenous bases (adenine, thymine, cytosine, guanine: adenine and quanine purines, thymine and cytosine pryimidines)
- nitrogenous bases paired by hydrogen bonds: adenine bonds by double hydrogen bond to thymine, cytosine bonds by triple hydrgoen bond to guanine

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8
Q

structure of DNA

deoxyribonucleic acid

A
  • double helix like twisted ladder, consist of 2 strands running in opposite direction (5’ to 3’ and 3’ to 5’)
  • polymer consisting of repeating units of nucleotides, these consist of** 5-carbon sugar (deoxyribose), a phosphate, and a nitrogen base**
  • carbon atoms in deoxyribose numbered 1 to 5
  • four nitrogenous bases (adenine, thymine, cytosine, guanine: adenine and quanine purines, thymine and cytosine pryimidines)
  • nitrogenous bases paired by hydrogen bonds: adenine bonds by double hydrogen bond to thymine, cytosine bonds by triple hydrgoen bond to guanine
  • gets packed and unpacked in nucleus as needed
  • eukryotic DNA combines with large amounts of histones (protein) from which it seperates briefly during replication
  • DNA + histones = chromatin
  • double helix of DNA wraps twice around core of histones, forming nucleosomes
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9
Q

structure of RNA

ribonucleic acid

A
  • single-stranded helix with repeating nucleotides: A, C, G, U (uracil which replaces T)
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10
Q

DNA replication in eukaryotes

A
  • double helix unzips, and each strand is a template for formation of new strand with complementary nucleotides
  • replication begins at sites called origins of replication where 2 strands seperate to form replication bubbles
  • 1000s of bubble seen along DNA molecule, and speeds up process of replication along giant DNA molecule with 6 billion nucleotides, bubbles expands as replication proceeds in both directions at once
  • replication fork at ends of replication bubbles (Y-shaped region where new strands of DNA elongating), all bubbles eventually fuse
  • enzyme DNA polymerase catalyzes antiparallel elongation of new DNA strands
  • DNA polymerase builds new strand from 5’ to 3’ direction by moving along template strand and pushing replication fork ahead (50 nucleotides/second elongation in humans)
  • DNA polymerase cannot initiate synthesis but can only add nucleotides to 3’ end of existing chain (consist of RNA and called RNA primer, enzyme primase makes primer by joining RNA nucleotides)
  • DNA polymerase replicates 2 original strands of DNA differently, even though it builds new strands in 5’ to 3’ direction, 1 strand formed towards replication fork in unbroken/linear fashion (leading strand)
  • lagging strand forms in direction away from replication fork in series of segments (Okazaki fragments) which are 100-200 nucleotides long + joined into continuous strand by enzyme DNA ligase
  • other proteins + enzymes assist in replication: helicases is enzyme that untwist double helix at replication fork which seperates 2 parental strands
  • single-stranded binding proteins is scaffolding, holding strands apart
  • topoisomerases lessen tension on tightly wound helix by breaking, swiveling, and rejoing DNA strands
  • DNA polymerases carry out mismatch repair (proofreading that corrects errors), damaged regions of DNA excised by DNA nuclease
  • everytime DNA replicates, some nucleotides from ends of chromosomes are lost, so special nonsense nucleotide sequences at ends of chromosomes that repeat thousands of times
  • protective ends called telomeres and are created and maintained by enzyme telomerase
  • telomeres get shorter everytime, clock that counts cell divisions and causes cell to stop dividing as cell ages

a cell can replicate entire DNA in a few hours

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11
Q

DNA to protein

A
  • triplet code in DNA transcribed into codon sequence in mRNA inside nucleus
  • newly formed strand of RNA (pre-RNA) then processed or modified in nucleus
  • codon sequence leaves nucleus and is translated into amino acid sequence (polypeptide) in cytoplasm at ribosome
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12
Q

3 RNA types involved in protein synthesis

A

3 RNA types directly involved in protein synthesis:
1. Messenger RNA (mRNA) in transcription: when sequence of DNA expressed, 1 strand is copied into mRNA according to base-pairing rules
2. Ribosomal RNA (rRNA) in translation: structural and makes up ribosome, which consists of 2 subunits (large + small) with 1 mRNA binding site and 3 tRNA binding sites (A, P, E)
3. Transfer RNA (tRNA) carries amino acid from cytoplasmic pool of amino acids to mRNA at ribosome: shaped like coverleaf and and has binding site for an amino acid at one end and another binding site for anticodon sequence that binds mRNA at the other

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13
Q

transcription

A

process by which information in DNA sequence copied into complementary RNA sequence

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13
Q

3 stages of transcription

A

initiation, elongation, termination

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14
Q

initiation

A
  • begins when enzyme, RNA polymerase recognizes and binds to DNA at promoter region
  • promoter tells RNA polymerase where to begin transcription and which two strands to transcribe
  • colllection of proteins (transcription factors) recognize key area within promoter (the TATA box, named for its repeating thymine and adenine nucleotides) and mediate binding of RNA polymerase to DNA
  • completed assembly of transcription factors and RNA polymerase bound to promoter called transcription initiation complex
  • one RNA polymerase attached to promoter, DNA transcription of DNA template begins
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15
Q

elongation

A
  • elongation of strand continues as RNA polymerase adds nucleotides to 3’ end of growing chain
  • RNA polymerase pries two strands apart and attaches RNA nucleotides according to base pairing rules
  • stretch of DNA transcribed into mRNA molecule is called a transcription unit
  • each unit consists of triplets of bases called codons that code for specific amino acids
  • single gene can be transcribed into mRNA simultaneously by several molecules of RNA polymerase following each other
  • has mechanisms for proofreading during transcription
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16
Q

termination

A

final stage, elongation continues for short distance after RNA polymerase transcribes termination sequence (AAUAAA), mRNA is cut free from DNA template

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17
Q

RNA processing

A

before pre-RNA strand shipped out of nucleus to ribosome, it is altered by enzymes:
- ** 5’ cap** (with a modified guanine nucleotide) added to 5’ end, helps RNA strand bind to ribosome during translation
- poly (A) tail (a string of adenine nucleotides) added to 3’ end, protects RNA strand from degradation by hyrolytic enzymes, and facilitates release of mRNA from nucleus into cytoplasm
- noncoding regions of mRNA called introns/intervening sequences spliced by SNPs (small nuclear ribonucleoproteins) within splicesomes, allows only exons (expressed sequences) to leave nucleus
- mRNA that leaves nucleus is a lot shorter than original transcription unit

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18
Q

alternative splicing

A

different RNA molecules produced from same primary transcript, depending on which RNA segments are exons vs. introns
regulatory proteins specific to cell type controls intron-exon choices by binding to regulatory sequences within primary transcript

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19
Q

translation of mRNA

A
  • process by which codons of mRNA sequence are changed into amino acid sequence
  • amino acids present in cytoplasm carried by tRNA molecules to codons of mRNA strand at ribosome according to baes-pairing rules
  • one end of tRNA w/ specific amino acid, and other end w/ anticodon (nucleotide triplet)
  • amino acid joined to correct tRNA by enzyme aminoacyl-tRNA synthetase
  • 20 different aminoacyl-tRNA synthetase (1 for each amino acid), 64 codons (61 coding for amino acids, AUG codes for amino acid methione and is also a start codon, UAA UGA and UAG are stop codons and terminate translation)
  • some tRNA molecules with anticodons to recognize 2+ different codons because pairing rules for third base of codon are not as strict as first 2 bases (relaxation known as wobble)

tRNA is used repeatedly unlike mRNA, energy provided by GTP (molecule similar to ATP)

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20
Q

initiation

in translation

A
  • begins when mRNA attached to subunit of ribosome
  • first codon is always AUG and must be positioned correctly for transcription of amino acid sequence to begin
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21
Q

elongation

in translation

A
  • continues as tRNA brings amino acids to ribosome and polypeptide chain is formed
  • a mRNA molecule translated simultaneously by several ribosomes in clusters (polyribosomes)
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22
Q

termination

in translation

A
  • termination of mRNA strand complete when ribosome reaches one of three stop codons
  • release factor breaks bond between tRNA and last amino acid of polypeptide chain
  • poly peptide freed from ribosome, and mRNA is broken down
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23
Q

The Genetic Code

A
  • 64 possible combinations of the 4 bases
  • there are redundancies in the code, but no ambiguity
  • code is universal and unifies all life, indicating that code originated early in evolution
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24
Q

gene mutation

A
  • permanent changes in genetic material which occur spontaneously and randomly
  • can be caused by mutagenic agents (chemicals/radiation)
  • mutations in somatic cells disrupts normal cell function
  • mutations in gametes transmitted to offspring and changes gene pool of population
  • mutations are the raw material for natural selection
  • some regions of DNA more vulnerable to mutation (A + T more breakages because of double bond rather than triple)
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25
Q

point mutation

A
  • simplest mutation
  • base-pair substitution (chemical change in 1 base pair in single gene)
  • sometimes can be beneficial or no change (wobble)
26
Q

insertion/deletion

A
  • single nucleotide insertion/deletion
  • results in frameshift
  • mutated/no polypeptide formed
27
Q

missense mutations

A

when point mutations/frameshifts change codon within gene to stop codon, translation is altered into missense/nonsense mutation

28
Q

genetics of viruses

A
  • parasite that can live only inside another cell
  • commandeers host cell machinery to transcribe/translate all proteins needed for new viruses
  • thousands new viruses formed and host cell often destroyed
  • DNA or RNA enclosed in protein coat (capsid)
  • some have viral envelope derived from membranes of host cells, cloaks capsid, and helps virus infect host
  • 1 virus type can only infect 1 cell type because it enters cell by binding to specific receptors on surface
  • 1 virus usually only infects 1 species, range of organisms a virus can attack is the host range
29
Q

bacteriophages or phage virus

A
  • most compex + best understood virus that infects bacteria
  • can reproduce in different ways:
    1. in lytic cycle, enters host cell, takes control of machinery, replicates, causes cell to burst, release new generation of phage viruses which repeats the cycle (virulent phage)
    2. in lysogenic cycle, replicates without destroying host cell, becomes incorporated into specific site in DNA, and remains dormant within host genome (prophage)
    when host cell divides, phage is also replicated, eventually environment trigger causes prophage to switch to lytic phase: viruses capable of both modes are temperate viruses
30
Q

retroviruses

A
  • contains RNA instead of DNA
  • after infecting host cell, retrovirus RNA is template for synthesis of complementary DNA (cDNA), reverses usual flow of information from DNA to RNA
  • reverse transcription occurs under direction of enzyme reverse transcriptase
  • usually inserts itself into host genome, becomes permanent resident (prophage), and makes multiple copies of viral genome for years
31
Q

transduction

A
  • phage viruses get bits of bacterial DNA as they infect cells which leads to genetic recombination (tranduction)
  • two types of tranduction: (generalized/restricted)
  • generalized moves random pieces of bacterial DNA as phage lyses one cell and infects another
  • restricted involves transfer of specific pieces of DNA (during lysogenic cycle, phage integrates into host cell at speicfic site and will sometimes carry adjacent host DNA with it and insert into next host)
32
Q

genetics of bacteria

A
  • bacterial chromosome: circular, double-stranded DNA tightly condensed into structure with small amount of protein, located in nucleoid region without membrane
  • replicates DNA in both directions from single point of origin
  • can reproduce w/ conjugation or binary fission (main method, asexual)
  • binary fission results with identical genes, but mutations occur
33
Q

bacterial transformation

A
  • Frederick Griffith (1927)
  • natural or artificial process for recombination of genetic information: small pieces of extra-cellular DNA taken up by bacterium leading to stable genetic change in recipient
34
Q

plasmid

A
  • foreign, small, circular, self-replicating DNA molecule that inhabits bacterium
  • bacterium can take in many, and will express genes carried by plasmid
35
Q

the operon

A
  • discovered in E. coli by Jacob and Monod (1940s)
  • important model of gene regulation in bacteria
  • set of genes and switches that control expression of genes
  • two types: inducible (lac) and repressible (tryptophan)
36
Q

tryptophan operon

A
  • promoter and 5 adjacent structural genes (A, B, C, D, E) that codes for 5 enzymes necessary to synthesize amino acid tryptophan
  • if RNA polymerase binds to promoter, one long strand of mRNA containing start and stop codons is transcribed
  • if enough tryptophan is present, tryptophan itself acts as **corepressor ** activating repressor
  • activated repressor binds to operator preventing RNA polymerase from binding to promoter
  • is a repressible operon, will always be on unless repressor is activated
37
Q

lac operon

A
  • E.coli needs lactose as energy source, so 3 enzymes must be syntheized to break down lactose into glucose and galactose
  • enzymes coded for by 3 genes in lac operon (A, B, C)
  • to be transcribed, repressor must be prevented from binding to operator and RNA polymerase must bind to promoter region
  • allolactose (isomer of lactose) is inducer that facilitates process by binding to active repressor and inactivating it
38
Q

CAP and cAMP - positive gene regulation

A
  • when glucose and lactose both present, E. Coli will first metabolize glucose, if lactose is present and glucose is short on supply, it will switch
  • ability to switch is from allosteric regulatory protein (CAP and cAMP)
  • attachment of CAP to promoter directly stiumlates gene expression = positive gene regulation
39
Q

RNA polymerase

A

enzyme that transcribes new RNA chain by linking ribonucleotides to nucleotides on DNA template

40
Q

operator

A

sequence of nucleotides near start of operon for active repressor to attach, binding of repressor prevents RNA polymerase from attaching to promoter and transcribing operon’s genes

41
Q

promoter

A

nucleotide sequence in DNA of gene that is binding site of RNA polymerase, positioning RNA polymerase to begin to transcribe RNA at appropriate position

42
Q

repressor

A

protein that inhibits gene transcription (binds to operator in operon)

43
Q

regulator gene

A

gene that codes for repressor, located away from operon and has own promoter

44
Q

prions

A
  • not cells nor viruses
  • misfolded versions of protein normally found in brain
  • if prion gets into normal brain, normal versions of protein will misfold the same way
45
Q

the human genome

A
  • 3 billion base pairs of DNA, about 24 000 genes
  • tiny fraction of DNA codes for proteins, much of it still gets transcribed into RNA
  • nongene DNA includes regulatory and repetitive sequences
46
Q

regulation of gene expression

A

cell expresses small precentage of genes at a time, expression of genes tightly regulated

47
Q

regulation at chromatin structure level

A
  • eukaryotic DNA packaged with histones (proteins) into chromatin, basic unit of the nucleosome
  • changes to histone structure changes chromatin configuration, binding more tightly/loosely making DNA less or more accessible for transcription/expression
  • acetylation of histone tails promotes loosening of structure and permits transcription
48
Q

regulation by methylation

A
  • methylation of certain bases (adding methyl groups) will silence DNA temporarily
  • removing methyl groups from methylated regions can turn genes on
  • responsible for long-term X-chromosome deactivation in females + long term deactivation of genes necessary for normal cell differentiation in embryonic development
49
Q

epigenetic inheritance

A
  • alterations to the genome that do not directly involve nucleotide sequence
  • not sure about mechanism, but environmental factors can alter expression of genes

reversible unlike mutations

50
Q

regulation at transcription level

A
  • transcription highly regulated
  • to initiate transcription: RNA polymerase must bind to promoter and requires transcription factors
51
Q

regulation at post-transcriptional regulation level

A
  • alternative RNA splicing is important way of regulating gene expression in which mRNA molecules produced from same primary transcript (dependong on which segments are introns vs. exons)
  • regulatory proteins specific to cell type will control intron-exon choices by binding to RNA sequences within primary transcript

90% of human-protein genes subject to aternative splicing

52
Q

degradation of mRNA

A

span of time before degradation of mRNA also regulates gene expression

rapid degradation of mRNA in bacteria makes it more adaptable to changes in environment
human mRNA continually translate protein for hours to weeks

53
Q

ncRNA

A
  • 90% of non-protein-coding DNA is transcribed into noncoding RNA (ncRNA)
  • ncRNAs bind to and assisted by specialized binding proteins (Argonaute) proteins
  • noncoding regions are not useless, and regulate our DNA
54
Q

microRNA (miRNA)

A
  • single-stranded RNA, 22 nucleotides long, forms complex with proteins
  • targets specific mRNA molecules to degrade or block translation
55
Q

small interfering RNA (siRNA)

A
  • similar to miRNA in size and function
  • blocking of gene expression by siRNA called RNA interference (RNAi)
56
Q

piwi-associated RNA (piRNA)

A
  • large class of ncRNAs that guide PIKWI proteins to complementary RNAs which are derived from transposable elements
  • protect germ line cells from attack by transposons
57
Q

proteins modified at post-translation level

A
  • newly made protein may spontaneously fold into correct shape and function
  • other newly made proteins must be activated before functioning
58
Q

recombinant DNA

A
  • taking DNA from 2+ sources and combining them into 1 molecule
  • naturally occurs in viral transduction, bacterial transformation, conjugation, transposons
  • genes also manipulated/engineered in labs (biotech or genetic engineering)
  • used to produce protein product (e.g. insulin), replace nonfunctioning gene in person’s cell with functioning gene by gene therapy, prepare multiple copies of a gene for analysis, engineer bacteria to clean up environment
59
Q

techniques of gene cloning

A
  • isolate gene of interest
  • insert gene into plasmid
  • insert plasmid into vector (cell that will carry plasmid)
  • clone the gene (bacteria reproduce by fission, plasmid and selected gene also cloned)
  • identify bacteria with gene and harvest it
60
Q

restriction enzymes

A
  • basic biotech tool from late 1960s
  • extracted from bacteria, which use them to defend against phages
  • cuts DNA at specific recognition sequences/sites
  • cuts are staggered and leaves single-stranded sticky ends to form temporary union with other sticky ends
  • fragments from cuts are restriction fragments
61
Q

gel electrophoresis

A
  • seperates large DNA molecules on basis of their rate of movement through agarose gel in electric field (DNA which is negative will flow from cathode (-) to anode (+))
  • concentration of gel altered to provide greater impediment to DNA, allowing for finer seperation
  • used to seperate proteins and amino acids
  • DNA must be cut by restriction enzymes before running in gel
  • seperated DNA can be analyzed in many ways
  • DNA strands can be sequenced to determine sequence of bases
  • DNA probe can identify location of specific sequence within DNA
62
Q

DNA probe

A
  • **radioactively labeled single strand ** of nucleic acid molecule to tag sequence in DNA sample
  • bonds to complementary sequence anywhere it occurs
63
Q

polymerase chain reactions (PCR)

A
  • made in 1985
  • cell-free, automated technique where piece of DNA can be rapidly copied/amplified
  • billions of copies of a fragment of DNA produced in a few hours
  • DNA piece to be amplified placed into test tube with Taq polymerase along with supply of nucleotides and primers necessary for DNA synthesis
  • amplified DNA to be stuided or compared with other samples
  • limitations: some information about nucletoide sequence in target DNA must be known before to make necessary primers, size of target piece must be very short, contamination is big problem
64
Q

restriction fragment length polymorphism (RFLPs)

A
  • restriction fragment is segment of DNA from restriction enzymes
  • restriction fragment pattern from noncoding regions (junk DNA) in human DNA is different in every individual
  • everyone’s RFLPs are unique (exception of twins), and inherited in Mendelian way - used accurately in paternity suits to determine father of a child
65
Q

complementary DNA (cDNA)

A
  • when trying to clone human gene in bacterium, bacteria lack introns and can’t edit them out after transcription
  • to clone human gene in bacterium, gene inserted must have no introns
  • scientists extract fully processed mRNA from cells and use reverse transcriptase to make DNA transcripts of RNA
  • resulting DNA molecule carries complete coding sequence without introns
  • DNA produced by retroviruses is complementary DNA (cDNA)
  • CRISPR (clustered regularly interspersed short palindromic repeats) is genetic engineering tool made of RNA that can be guided by enzyme CAS9 to modify stretch of DNA, targets particular sequence and permanently disrupts/add small pieces of corrective DNA