(MQ) Unit 6: Gene Expression and Regulation

Identify an example of when a PCR should be done.

The cloning of gene fragments to analyze genetic diseases, identification of contaminant foreign DNA in a sample, and the amplification of DNA for sequencing

How do siRNA and miRNA affect gene expression?

siMRAs inhibit the expression of one specific target mRNA, while miRNA's regulate the expression of multiple mRNAs

Which strand is the template strand?

3' to 5' strand

Which direction is the template strand READ?

3-5

If there is 20% thymine in a DNA strand, how much cytosine is there?

20% T means 20% A; So G and C = 60% Cytosine = 60/2 = 30%

How many nucleotides make up a codon?

3

Identify three post-transcriptional modifications that alter the pre-mRNA prior to its release from the nucleus.

5' cap is added to the 5' end to protect it from exonucleuses which target 5' ends A poly-A tail is added to the 3' end of the RNA molecule to help the RNA molecule leave the nucleus and to protect it from exonucleases Noncoding introns are removed from the RNA strand and the exons are spliced together

What direction is DNA synthesized?

5' to 3'

Which direction does transcription take place (which direction is the RNA synthesized)?

5' → 3'

What is the TATA box?

A DNA sequence in eukaryotic promotors crucial in forming the transcription initiation complex

What is a repressor?

A DNA- or RNA-binding protein that inhibits the expression of one or more genes by binding to the operator or associated silencers

What are the nucleotide base pairing rules?

A bonds with T; G bonds with C (A bonds with U in RNA)

What is gel electrophoresis?

A laboratory method used to separate mixtures of DNA, RNA, or proteins according to molecular size (largest pieces travel farthest down because of polar attraction)

What is transformation?

A process in which a prokaryote takes in DNA found in its environment that is shed by other prokaryotes, thereby changing morphology and physiology

What is bacterial transformation?

A process in which foreign DNA is taken up from the environment by a cell

Describe the trp operon.

A repressible operon, transcription is usually on, but can be inhibited when tryptophan binds allosterically to the regulatory protein (positive gene regulation - changes shape of repressor to allow it to bind to operator and stop transcription)

How does the repressor interact with the promoter?

A repressor is a protein that turns off the expression of one or more genes. The repressor protein works by binding to the gene's promoter region, preventing the production of messenger RNA (mRNA).

Negative Gene Regulation

A repressor protein binds to an operator to prevent a gene from being expressed

What is a promoter?

A sequence of DNA to which proteins bind that initiate transcription of a single RNA from the DNA downstream of it. This RNA may encode a protein, or can have a function in and of itself, such as tRNA, mRNA, or rRNA

What is a codon?

A sequence of three nucleotides that together form a unit of genetic code in a DNA or RNA molecule (one amino acid)

What is a plasmid?

A small, double stranded extrachromosomal DNA molecule often carrying genes for antibiotic resistance and found in bacteria and sometimes present in archaea and eukaryotic organisms

What is the promoter region?

A specific sequence of DNA bases at the start of a gene on the sense strand where RNA polymerase binds (defines start of transcription)

What is an operon?

A unit made up of linked genes that is thought to regulate other genes responsible for protein synthesis. A sequence of DNA containing a cluster of genes under the control of a single promoter

What codons code for lysine?

AAA, AAG

How is DNA modified to initiate transcription (to regulate gene expression)?

Activator transcription factors attach to the promoter and recruit RNA polymerase to increase transcription

What occurs during a bacterial transformation?

Addition of CaCl2 and heat shocking are used to induce bacterial cells to pick up recombinant plasmids and express foreign DNA

Which of the nitrogenous bases are purines?

Adenine and Guanine

How does the genetic code demonstrate common ancestry?

All living organisms store their genetic information using the same nucleotide bases

How can multiple proteins be synthesized from the same mRNA transcript?

Alternative Splicing (cutting out different parts in pre-mRNA stage)

What happens in the elongation step of translation?

Amino acids are brought to the ribosome by tRNAs and linked together to form a chain, the methionine-carrying tRNA starts out in the middle slot of the ribosome, called the P site. Next to it, a fresh codon is exposed in another slot, called the A site. The A site will be the "landing site" for the next tRNA, one whose anticodon is a perfect (complementary) match for the exposed codon.

Inducible vs. Repressible Operons

An inducible operon is able to be turned on (positive control) and a repressible operon is able to be turned off (negative control) Inducible - Protein allows repressor to fall off operator Repressible - Protein allow repressor to bind to operator

Describe how incorrect chromosome numbers result in human disorders.

Aneuploidy: monosomy-1 chromosome, trisomy- 3 chromosomes Having incorrect numbers leads to changes, depending on what the chromosome was responsible for

How does horizontal transfer increase variation?

By adding new material to the gene pool, phenotypes will change

How does a retrovirus violate the central dogma?

By synthesizing DNA from their RNA genomes, and also by replicating their RNA genomes.

What is the function of the poly-A tail?

Helps mRNA leave the nucleus, protects it from degradation (stability)

What happens if an insertion or deletion occurs at a nucleotide base pair? How does this affect the protein product? How does this affect the amount of the protein product?

Can cause a frameshift; all nucleotides downstream will be improperly grouped into codons, affecting protein production; missense mutations caused by the frameshift can cause nonsense mutation and a premature termination - protein will be (mostly) non-functional

How do mutations affect gene products?

Can introduce new alleles which increase genetic variety = proteins changes

How can a mutation have a negative effect on the product produced?

Cause proteins to not form correctly, proteins can't function in the same way, causing genetic disorders or cancer

How is prokaryotic DNA shaped?

Circular

What reproductive processes increase genetic variation?

Crossing over segregation independent assortment random fertilization

What is the function of RNA splicing?

Cutting out the introns (non-coding) with spliceosomes and splicing the exons (coding) back together to prepare for translation

What are epigenetic changes?

Modify the activation of certain genes, but not the genetic code sequence of DNA (changes in gene expression)

What occurs during a PCR?

DENATURING: DNA sample is heated so double stranded DNA separates ANNEALING- Sample is cooled allowing DNA primers bind to target sequences on DNA ELONGATION (EXTENSION): "Taq polymerase" enzyme synthesizes two new strands of DNA, using the original strands as templates. The cycle of heating and cooling repeated as many as 30 or 40 times, leading to more than one billion exact copies of the original DNA segment.

What is the primary source of heritable information?

DNA

What is the central dogma?

DNA -> RNA -> Protein

What information can be drawn from a gel electrophoresis?

DNA comparisons can identify differences in individual's DNA (used in crime)

Why is DNA a better hereditary material than RNA?

DNA is chemically more stable than RNA. RNA has a hydroxyl group (-OH group) in it which leads to RNA being lesser stable

What is the function of DNA ligase?

DNA ligase joins the Okazaki fragments on the lagging strand together.

How is DNA modified to inhibit transcription (to regulate gene expression)?

DNA methylation - methyl groups are added to DNA which tightly packs it to reduce transcription; repressor transcription factors bind to the promoter to block RNA polymerase from attaching to the DNA

What is the function of DNA polymerase?

DNA polymerase can only add on to the 3' end of an existing nucleic acid strand. DNA polymerase is responsible for creating and expanding the new strands of DNA. Once the DNA polymerase is attached to the origin site of the DNA (Template strand) it can start making the new DNA by bringing in free floating nucleotides and creating the complementary strand. The DNA polymerase can extend the primer by adding nucleotides only to the 3' end.

What direction does the DNA polymerase READ the DNA template?

DNA polymerase requires a free 3' OH group for initiation of synthesize; it has to extend in one direction by extending the 3' end of the preexisting chain. So the DNA polymerase moves along the template strand in a 3'-5' direction and the daughter strand is formed in a 5'-3' direction.

What occurs during gel electrophoresis?

DNA separates (DNA is negative and moves towards the positive electrode from the negative electrode) according to size; the smallest fragments will be farthest from the well (where the DNA was loaded), and the heavier molecular weight fragments will be closest to the well.

What is DNA?

Deoxyribonucleic acid: it stores all the genetic information containing heredity information to make all proteins, and traits.

How can differential gene expression affect the cellular products?

Depending on the cell type there can be different activators available to control which genes are expressed in which tissues; this will cause specific products to be produced only in specific tissues (example: albumin gene is expressed in liver cells, but not in lens cells)

How can differential gene expression affect the cellular functions?

Depending on which genes are expressed in which tissues, specific proteins will be made leading to more specific cellular functions

Why can different phenotypes result from different levels of gene expression?

Different tissues will express different genes based on which activators/transcription factors are present in those cell types. Alternative splicing in different cell types can also result in different genes being transcribed and translated, causing different proteins to be expressed.

How is the amount of DNA different between a prokaryote and a eukaryote?

Eukaryotes have much more DNA than prokaryotes

Identify two functions of plasmids.

F plasmids contain genes that code for the production of sex pili and enzymes required for conjugation. R plasmids contain genes for antibiotic resistance

True or False: One codon can code for more than one amino acid.

False

True or False: Natural selection acts on genotype.

False (acts on physical conditions of organisms)

What is transduction?

Foreign DNA is introduced into a cell by a viral infection

How is genetic information transmitted from one generation to the next?

Genes (DNA)

What is transposition?

Genes are transferred from one organism to another through the copying and inserting process. Transposition accounts for the antibiotic resistance and transmission of virulence in certain bacteria. The mobile DNA segment that is associated with transposition is called transposon or jumping genes.

How is genetic engineering used to manipulate DNA?

Genetic engineering is the process of using recombinant DNA technology to alter the genetic makeup of an organism; therefore, you are directly manipulating one or more genes to obtain a desired phenotype

How does a genetic change enhance survival?

Genetic variation is advantageous because it allows some individuals to adapt to the environment which will allow them to better survive

How does a change in genotype affect the phenotype?

Genotype determines phenotype

Identify two examples of retroviruses.

HIV, SIV (simian) and human T-cell lymphotropic virus type 1 (HTLV-1) causes a form of cancer called adult T-cell leukemia (ATL)

What does it mean if an organism is a triploid?

Has three sets of chromosomes

How are histones modified to initiate transcription (to regulate gene expression)?

Histone acetylation - removes the positive charge on the histones, and the condensed chromatin is transformed into a more relaxed structure (relaxed form)

How are histone modified to inhibit transcription (to regulate gene expression)?

Histone deacetylation - removes the negative charge on the histones, DNA becomes more tightly wrapped around the histones, making it harder for transcription factors to bind to the DNA (condensed form)

What chemical process releases the growing polypeptide?

Hydrolysis The release factors cause the ribosome peptidyl transferase to add a water molecule to the carboxyl end of the most recently added amino acid in the growing polypeptide chain attached to the P-site tRNA. This causes the polypeptide chain to detach from its tRNA, and the newly-made polypeptide is released.

What is the function of gel electrophoresis?

Identify differences in DNA samples

How can a mutation have a neutral effect on the product produced?

If they don't change the amino acid sequence, and it doesn't change the protein coding

How is DNA stored?

In eukaryotes it is in the nucleus coiled around histones or in the nucleoid region of prokaryotes

How is genetic engineering used to analyze DNA?

In many cases genetic engineering through PCR, gel electrophoresis, and recombinant DNA technology, will be used to determine how the gene on the DNA is expressed to cause a specific phenotype in that organism and how we can manipulate that expression in other organisms.

What is the function of bacterial transformation?

Incorporation of recombinant plasmids allows bacterial cells to express DNA from other organisms

Regarding the lac operon, is this operon repressible or inducible?

Inducible

Describe the process that takes place during transcription.

Initiation - RNA polymerase binds to the promoter sequence of DNA and then separates the DNA strands of the specific gene it is trying to transcribe. Elongation - the template strand of DNA acts as the template for RNA polymerase. RNA polymerase "reads" the template strand one base at a time and will add the complementary RNA nucleotides making a chain that grows from 5' to 3'. Termination - terminator sequences signal that the RNA transcript is complete. an example of a termination mechanism involves forming a hairpin in the RNA.

Describe the process of DNA replication using the enzymes described above (helicase, topoisomerase, DNA polymerase, primase, RNA primers, and DNA ligase)

Initiation: DNA replication is initiated at a specific coding region. This is where the replication fork is. There are multiple origin sites. DNA helicase enzymes will unwind the double helix and expose the two strands and the hydrogen bonds holding the two strands together will be broken. DNA primase will synthesize a small RNA primer which will act to start the process for DNA polymerase which is the enzyme responsible for creating and expanding the new strands of DNA. Elongation: Once the DNA polymerase is attached to the origin site of the DNA (Template strand) it can start making the new DNA by bringing in free floating nucleotides and creating the complementary strand. The DNA polymerase can extend the primer by adding nucleotides only to the 3' end. The new strand is formed in the 5' to 3' direction. The new strand is called the leading strand. The other template strand of the DNA is called the lagging strand (it is antiparallel) and is read in a 5' to 3' direction. Continuous DNA synthesis could have to be done in the 3' to 5' direction which is important because the bases would be added to the 5' end and we know they cannot do that. So this strand has to be created in fragments but still in the 5' to 3' direction and these fragments are called the okazaki fragment. Termination: Once there is no more DNA template to replicate or the two replication forks meet then the DNA replication can be terminated. RNAse H removes the RNA primer at the beginning of the okazaki fragment, and DNA ligase re joins the fragments together to create one complete strand and stabilizes the DNA.

How does a virus incorporate its viral genome into a host genome?

Integrase

What is the function of topoisomerase?

It breaks a covalent bond between deoxyribose sugar and a nitrogenous base in one parental strand temporarily during the replication process to make sure the DNA doesn't get twisted or coiled wrong.

What is the function of reverse transcriptase?

It catalyzes the formation of DNA from an RNA template (viral DNA)

What causes Turner syndrome?

It is caused by either a partial or complete loss (monosomy) of the second X chromosome (only has one X chromosome)

What is the function of transcription factors?

Mediate the binding of RNA polymerase and the initiation of transcription

Identify an example of when DNA sequencing should be done.

Medical diagnosis, forensics, biology of development and evolution; Comparing healthy and mutated DNA sequences can diagnose different cancers and can be used to guide patient treatment

Why is a triploid organism usually sterile?

Meiosis cannot proceed normally and instead produces mostly aneuploid daughter cells that cannot yield viable zygotes

How does the mRNA, tRNA, and rRNA all interact?

Messenger RNA (mRNA) carries the genetic information copied from DNA to the cytoplasm Transfer RNA (tRNA) is the key to deciphering the codons in mRNA. Each type of amino acid has its own type of tRNA, which binds it and carries it to the growing end of a polypeptide chain. The correct tRNA with its attached amino acid is selected at each step because each specific tRNA molecule contains a three-base sequence (anticodon) that can base-pair with its complementary codon in the mRNA. Ribosomal RNA (rRNA) associates with a set of proteins to form ribosomes. These complex structures, which physically move along an mRNA molecule, catalyze the assembly of amino acids into protein chains. They also bind tRNAs and various accessory molecules necessary for protein synthesis. Ribosomes are composed of a large and small subunit, each of which contains its own rRNA molecule or molecules.

What is the function of mRNA?

Messenger RNA carries copies of the genetic instructions from DNA to other parts of the cell.

What is involved in the cell's DNA repair mechanisms?

Mismatch repair: direct reversal of the damage, base or nucleotide excision repair, and postreplication repair (take out parts)

What is the function of the GTP cap?

Modified cap of 20-40 guanines added to the 5' end, this helps ribosomes attach to the 5' end once the mRNA reaches the cytoplasm

How is eukaryotic DNA shaped?

Multiple rod shaped chromosomes (linear)

What is the cause of new genetic variation?

Mutations

How do new phenotypes originate?

Mutations, horizontal gene transfer, recombination/ crossing over

What causes errors in DNA replication?

Mutations, insertions/deletions Can be caused by environmental factors like radiation

How can a mutation have a positive effect on the product produced?

New versions of proteins that are evolutionarily beneficial

What are the other names of the template strand?

Noncoding strand, antisense strand

Identify an error in meiosis that leads to a change in phenotype.

Nondisjunction (can result in XXY - a male with certain female traits)

What causes Down syndrome/Trisomy 21?

Nondisjunction at the 21st chromosome (trisomy = 3 chromosomes

Identify an error in mitosis that leads to a change in phenotype.

Nondisjunction or... Duplication repeats a segment. Inversion reverses a segment within a chromosome. Translocation moves a segment from one chromosome to another.

What occurs during the process of DNA sequencing?

One new sequencing technology involves watching DNA polymerase molecules as they copy DNA - the same molecules that make new copies of DNA in our cells - with a very fast movie camera and microscope, and incorporating different colors of bright dyes, one each for the letters A, T, C and G.

How do transcription factors affect the binding at the promoter?

Only after the transcription factors are attached to the promoter does RNA polymerase attach to it

Identify an example of when a gel electrophoresis should be done.

Paternity testing, determining evolutionary relationships, diagnosis of genetic diseases, identifying markers for inherited disorders (carriers)

How does natural selection affect phenotypes?

Phenotypes are a combination of genes and environmental influences, natural selection acts on the phenotype, which is the characteristics of the organisms which actually interact with the environment because a specific phenotype could provide a reproductive benefit and become more common in a population

How does the genotype influence the phenotype?

Phenotypic expression is the physical reflection of the possible gene combination

What is PCR?

Polymerase Chain Reaction; used to amplify DNA

What is the function of RNA primers and primase?

Primers are short strands of RNA and must be created (synthesized) by the enzyme primase. Primase is an RNA polymerase. The primers serve as a starting point for DNA synthesis to occur.

How does the location of translation affect gene expression in a prokaryote?

Prokaryotes can perform both transcription and translation simultaneously (eukaryotes have to remove mRNA from a nucleus in order to start transcribing)

How does the location of translation differ between a prokaryote and a eukaryote?

Prokaryotes only have free floating ribosomes; whereas eukaryotes have either free floating or bound ribosomes

What is the function of RNA polymerase?

RNA polymerase uses the DNA template strand to link RNA nucleotides together to form a complementary strand of RNA.

How are multiple codons able to code for the same amino acids?

Redundancy, not ambiguity (multiple codons code for the same amino acid; no codon codes for more than one amino acid)

What are regulatory sequences?

Regions that impact how genes are used (gene switches, promoters, leader/trailer, etc.) Segment of a nucleic acid molecule which is capable of increasing or decreasing the expression of specific genes within an organism

What is the enhancer region?

Regulatory DNA sequences that, when bound by specific proteins called transcription factors, enhance the transcription of an associated gene

Describe the lac operon.

Regulatory gene produces active repressor that binds operator and block rna pol. when lactose is available allolactose binds the repressor and inactivates it, so repressor falls off the operator. But low glucose levels means high cAMP levels, cAMP binds to CAP binding site on promoter and increases RNA polymerase efficiency. (repressor is removed when lactose is present, allowing digestive proteins to be made; negative gene regulation)

Positive Gene Regulation

Regulatory protein allows regulatory protein to bind to the operator and stop transcription. Substance present = no genes) (different from negative regulation, where regulatory protein removes regulator, allowing genes to be produced)

What information can be drawn from a bacterial transformation?

Reporter genes added to plasmids (EX: antibiotic resistance genes, GFP) can allow identification of cells that have incorporated the recombinant plasmid

Regarding the trp operon, is this operon repressible or inducible?

Repressible

Regarding the lac operon, what happens when lactose is absent?

Repressor active, operon off /glucose present, cAMP level low, little lac mRNA synthesized

Regarding the lac operon, what happens when lactose is present?

Repressor inactive, operon on /glucose scarce, cAMP level high, abundant lac mRNA synthesized

How do negative regulatory molecules inhibit gene expression?

Repressor protein binds to an operator to prevent a gene from being expressed

Where does translation take place?

Ribosomes in the cytoplasm

What are the three steps of translation?

Same as Transcription • Initiation • Elongation • Termination

What are the three types of substitution point mutations? How does each affect the protein product?

Silent, missense, and nonsense. silent: no change in amino acids missense: substitute another amino acid nonsense: early termination silent: no change missense: can have no effect or render the protein nonfunctional nonsense: protein shorter or unfinished

Why are RNA primers required for DNA replication?

Since DNA polymerase can only attach new DNA nucleotides to existing strands of nucleotides the primers serve as a foundation for DNA synthesis.

What information can be drawn from DNA sequencing?

Since the DNA sequence confers information that the cell uses to make RNA molecules and proteins, establishing the sequence of DNA is key for understanding how genomes work How related organisms are

What is miRNA?

Single-stranded miRNA (MicroRNA) are small, highly conserved non-coding RNA molecules involved in the regulation of gene expression by degrading their target mRNAs and/or inhibiting their translation.

What is siRNA?

Small interfering RNA. They block expression of a gene with the same sequence as the RNA (double stranded)

Where does translation start?

Start codon (AUG)

How does the DNA sequence determine the RNA sequence?

The RNA sequence will be the "opposite" of the DNA sequence, so that when it is transcribed, the original DNA message is used by tRNAs

What is differential gene expression?

The expression of different genes by cells with the same genome

What happens in the termination step of translation?

The finished polypeptide is released to go and do its job in the cell, termination happens when a stop codon in the mRNA (UAA, UAG, or UGA) enters the A site.

How does natural selection affect genotypes?

The genotype determines the phenotype, so if the phenotype is connected to the genotype due to a genetic basis, and a specific phenotype is better suited for the environment then that genotype will become more common in the population

What is horizontal gene transfer?

The introduction of genetic material from one species to another unrelated species by mechanisms other than the vertical transmission from parent(s) to offspring

How does the leading and the lagging strand differ?

The leading strand is synthesized in the 5'-3' direction and the lagging strand in the 3'-5' direction. The leading strand is created continuously while the lagging strand is created in fragments (the okazaki fragments).

What is DNA sequencing?

The process of determining the precise order of nucleotides within a DNA molecule

What is transcription?

The process of making RNA from DNA

What is cell differentiation?

The process of specialization leading to the formation of brain, heart, and all other tissues from the single-cell zygote and the primitive, three-layered embryo

What happens in the initiation step of translation?

The ribosome gets together with the mRNA and the "initiator" tRNA, carrying the first amino acid in the protein, which is almost always methionine (Met)

What is the function of helicase?

They separate the double helix by breaking the hydrogen bonding that is keeping the DNA molecule together so each strand can get copied.

What is the function of the promoter?

To initiate transcription (it's where the RNA polymerase attaches to on the DNA sequence)

What is the function of tRNA?

Transfer RNA transfers each amino acid to the ribosome as it is specified by the coded messages in the mRNA.

True or False: All somatic cells have the same DNA.

True

Regarding the trp operon, what happens when tryptophan is absent?

Tryptophan absent, repressor inactive, operon on

Regarding the trp operon, what happens when tryptophan is present?

Tryptophan present, repressor active, operon off (does not need to produce any when it is already present)

Why does a purine always pair with a pyrimidine?

Two purines bonding together would take up too much space between the two DNA strands, which would affect the structure and not allow the strands to be held together properly.

What is conjugation?

Type of prokaryotic reproduction, some bacteria exchange genetic material by pilus, DNA transferred can be in the form of a plasmid or as a hybrid, containing both plasmid and chromosomal DNA.

What amino acid is coded by UAU?

Tyrosine

Identify an example of when a bacterial transformation should be done.

Use of bacteria that have incorporated recombinant plasmids containing human genes to produce human products (somatostatin hormone, insulin)

Identify two biotechnical uses for plasmids.

Used as a tool to insert genes into bacteria to produce therapeutic proteins such as human insulin Cloning PCR (making large amounts of DNA copies)

What is the function of DNA sequencing?

Used to determine the exact sequence of bases (A, C, G, and T) in a DNA molecule.

What is the function of PCR?

Used to make copies of DNA fragments, increasing the size of the sample

How does a virus form progeny viruses?

Using the host cell's machinery the viral DNA will be transcribed and translated into viral proteins that will then be assembled with the copies of the viral genetic material.

How do triploids form?

When a haploid cell is fertilized by a diploid cell

How do viruses recombine genetic information?

When two different strains of a virus infect the same host cell, their genetic material can mix and produce a new, recombinant strain.

Which of the nitrogenous bases are pyrimidines?

cytosine, thymine, uracil

What is the structure of a purine?

double ring

What are the three sites found on a ribosome and what is their function?

e site (exit site) - discharges the tRNA p site (peptidyl- tRNA binding site) - holds the tRNA carrying the polypeptide a site (aminoacyl-tRNA binding site) - holds the tRNA carrying the next amino acid to be added to the polypeptide chain

What is the function of rRNA?

rRNA molecules help hold ribosomal proteins in place and help locate the beginning and end of the mRNA message. They may also carry out the chemical reaction that joins amino acids together.

What is the structure of a pyrimidine?

single ring