Genetics Final

Gene

(unit of inheritance) is a discrete piece of information (DNA) that encodes a functional product (RNA or protein) which determines our traits

Chromosomes

where our genes are located

Alleles

different forms of genes that may affect the encoded product

Genomes

complete set of genetic instructions

Phenotype

the outward appearance of a trait as affected by the genotype

Genotype

the allelic make-up of an individual

Explain how genes are connected to phenotype. Summarize the central dogma.

-genes encode for an organism’s genotype which determines the physical characteristics/phenotype

-the central dogma states that information is transferred from one DNA molecule to another which then gets transcribed into RNA and then translated into an amino acid sequence; in simpler terms DNA to RNA (transcription) to sequence (translation)

Describe the chemical composition and structure of nucleotides.

each nucleotide contains one phosphate group, a 5-carbon sugar, and a nitrogenous base

Explain how nucleotides assemble to form nucleic acids.

-nucleotides are catalyzed by DNA or RNA poly and connect to each other via covalent phosphodiester bonds

-on the 5’ end, is the phosphate group and 3’ end is the pentose sugar, keep in mind that the polymer is directional

Describe the structure of DNA and the importance of base pairing in DNA structure and function.

-DNA is a double helix structure composed of two antiparallel, complementary strands

-A always pairs with T (connected with 2 hydrogen bonds) and C always pairs with G (connected with 3 hydrogen bonds)

-the predominant form in cells is the B form, which is a loose, right-handed helix

What are the three DNA sequence types in eukaryotes?

unique sequences, moderately repetitive, highly repetitive/satellite

Unique Sequence DNA

single-copy genes with gene families, includes coding and noncoding DNA

Moderately Repetitive DNA

(150-300 bp, 1000s of copies) includes tandem repeats and interspersed repeats

Highly Repetitive/Satellite DNA

(10 bp or less) often in tandem repeats with millions of copies, and serves a structural role

Compare the structure and organization of prokaryotic vs eukaryotic chromosomes.

-eukaryotic chromosomes are linear while prokaryotic chromosomes are circular

-in prokaryotes, the chromosome is confined to the “nucleoid” with DNA associated with a variety of packaging proteins and contains plasmids (small, circular extrachromosomal DNA molecules) whereas in eukaryotes, chromosomes are located inside the nucleus and their structure changes with the cell cycle (G1, S, G2, M)

-in both bacteria and eukaryotes, DNA folds into loops to supercoil and obviously chromosomes contain the DNA

Compare the packaging mechanisms of prokaryotic vs eukaryotic chromosomes.

-eukaryotic DNA packages itself into chromatin and the major packaging proteins involved for eukaryotic DNA packaging are histones (dr cline’s fave)

-prokaryotic DNA package their chromosomes with nucleoid associated proteins

Contrast euchromatin and heterochromatin in eukaryotes.

-euchromatin is less condensed, on chromosome arms, has many unique sequences, replicated throughout S phase, is transcribed often, and crossing over is common

-heterochromatin is more condensed, located at centromeres, telomeres, and other specific places, has repealed sequences, few genes, is replicated late into the S phase, infrequently transcribed, and crossing over is uncommon

List the three components required for transcription.

-DNA template: to specify new RNA sequence

-Ribonucleotide subunits: assemble into new RNA molecule

-RNA polymerase: to catalyze synthesis of RNA

Summarize the differences between RNA and DNA chemistry and structure.

-DNA: composed of nucleotides, deoxyribose, (A,G,C,T), nucleotides joined by phosphodiester bonds, double stranded, double helix, stable

-RNA: composed of nucleotides, ribose, presence of 2’-OH group, (A,G,C,U), nucleotides joined by phosphodiester bonds, single stranded, many structures, easily degraded

Identify the three main types of RNA synthesized in both prokaryotes and eukaryotes, and the function of each.

-mRNA → carries genetic code for proteins

-rRNA → structural/functional components of the ribosome

-tRNA → helps incorporate amino acids into polypeptide chain

Differentiate between coding and template strands

-Template: read 3’ - 5’

-Coding: nontemplate strand is not transcribed; identical to the mRNA except for the U-T replacement

—RNA synthesis is complementary and antiparallel to the template strand

—New nucleotides are added to the 3’-OH group of the growing RNA so transcription proceeds in a 5’ →3’ direction

Promoter

where RNA pol is positioned at the beginning of the gene

Coding Region

DNA sequence that will be copied into RNA

Terminator

site that signals where transcription ends

Upstream/Downstream

positions relative to or within the gene

Describe the events and components associated with each of the three stages of bacterial transcription.

-Initiation, Elongation, termination (Rho-dependent or independent)

Steps:

-Sigma subunit directs RNA pol to promoter

-DNA unwound around start site

-Sigma subunit released after 10 NTPs

-Transition to elongation

-RNA pol rewinds DNA after synthesis

-Termination site encountered

-Dissociation of RNA pol, RNA, and template

Comparison of euk transcription and prokaryotic transcription

-basics are like prokaryote transcription

Differences:

-More than 1 type of RNA pol (RNA pol II)

-Promoter structure and sequence

-Mechanism of RNA pol recruitment to promoter

-Processing of RNA is common

Core Promoter

necessary but not sufficient for transcription; requires regulatory promoter (i.e. TATA box)

Regulatory Promoter

controls levels of transcription; where proteins that increase or decrease transcription will bind

General Transcription Factors

assemble on the core promoter, recruit RNA pol II (take role of sigma factor for euk transcription) (TFII)

Activators + Repressors bind where?

will bind to regulatory promoter or enhancers

Mediator

a large, crucial protein assembly in eukaryotes that acts as a bridge, relaying signals from DNA-binding transcription factors (activators/repressors) to RNA polymerase II (Pol II) and the general transcription machinery (GTFs) to control gene expression.

Coactivator

essential proteins that bridge DNA-bound activators (transcription factors) to the general transcription machinery (like RNA Polymerase II), helping to activate gene expression by modifying chromatin structure (e.g., histone acetylation/deubiquitylation) and facilitating assembly, essentially making genes accessible and boosting transcription initiation and elongation

Compare the structure of eukaryotic vs prokaryotic genes.

Prokaryotic genes are simple, continuous coding sequences on a single, circular DNA in the cytoplasm, lacking introns and histones; eukaryotic genes are complex, split into coding exons and non-coding introns, located on multiple, linear chromosomes within the nucleus, wrapped around histone proteins.

Summarize the three main types of processing that occur for pre-mRNA, and the purpose of each

-5′ cap: modified G nucleotide, protects RNA, required for ribosome binding

-Splicing: removal of introns, joining of exons

-Poly A tail: added to 3′ end, stabilizes RNA, aids in export and translation

Describe how nuclear pre-mRNA splicing is accomplished, including how splice sites are identified

-Complex of 5 small nuclear RNAs and nearly 300 proteins

-Each snRNA associates with certain proteins to formsnRNP - “snurp”

-RNA components recognize splice sites and catalyze splicing

-Produces “lariats” of spliced out introns and mRNA

Explain the significance of alternative splicing.

-Some exons are removed during splicing

-Yields alternative series of exons and thus a different mRNA sequences

-This allows one gene to code for many different related proteins

-Often occurs for 95% of genes in humans with multiple exons

-It is often used to generate similar proteins for expression in different tissues

Summarize the roles of mRNA, tRNA, and rRNA in the process of translation

-mRNA provides template

-trnas read template and deliver template-specified amino acids

-rrnas and proteins form the translation machinery (the ribosome)

Compare and contrast prokaryotic vs. eukaryotic mRNA.

Both read in the 5’ to 3’ direction and contain untranslated regions. However, eukaryotic mRNA is processed and is monocistronic whereas most prokaryotic mRNA is polycistronic.

Use the genetic code to predict the amino acid sequence coded by a given RNA sequence

Use the chart to remember that three-nucleotide sequences, known as codons, code for one amino acid each (although some codons can code for the same amino acid; “wobble”) and U = T.

Explain the structure of tRNA and how that structure is critical to its function.  Describe how aminoacyl tRNA synthetase is critical for tRNA function.

tRNA has a “cloverleaf” structure via internal complementary base pairing. Which serves as an adaptor between nucleotides and amino acid sequences. tRNA needs to carry the correct anticodon; needs to be “loaded” with the correct amino acid which is carried out by amino-acyl tRNA synthetases. The aminoacyl-tRNA enzymes are specific to each AA, so there is a different synthetase for the amino acid.

Contrast how the start codon is identified in prokaryotes vs eukaryotes

Prokaryotic: 16s ribosomal subunit identifies the Shine-Delgarno sequence to bind to the start codon.

Eukaryotic: 40s ribosomal subunit binds to the 5’ GTP cap and will scan to find the start codon which is typically near the Kozak sequence (ANNAUGG).

Summarize the key events in each of the three stages of translation

Initiation: small ribosomal subunit binds and searches for AUG, binds, translation factors disassociate. Large subunit binds, tRNA starts to bring amino acids; start of transition to elongation.

Elongation: tRNAs start to bring AAs to their respective codons, peptide bonds are catalyzed by rRNA of large subunit, large and small subunits move one codon in the 3’ direction; protein synthesis is N-terminus to C-terminus.

Termination: elongation continues until the stop codon is translocated into A site. Release factor recognizes stop codon, ribosome moves one codon in 3’ direction and machine disassembles.

Constitutive Expression

genes that are expressed on a continual basis “housekeeping genes”

Regulated Expression

genes expressed only when the product is needed

Structural Genes

code for proteins involved in metabolism or provide a structural function in the cell.

Regulatory Genes

code for proteins/RNA that interact with other DNA sequences and affect the expression of those sequences

Describe how gene expression may be regulated at different levels

Alteration of chromosome structure, initiation of transcription (which depends on the interaction of proteins or RNA molecules with DNA regulatory elements), mRNA processing, mRNA stability, translation, posttranslational modification.

Inducible

usually “off” only activated when required by cell

Repressible

usually “on” inactivated when not required by cell

Positive Control

regulator protein binds & activates transcription (activator)

Negative Control

regulator protein binds & inactivates transcription (repressor)

Compare and contrast gene regulation in eukaryotes with that of prokaryotes.

In eukaryotes, gene control is very important whereas it is absent in prokaryotes. In eukaryotes, initiation of transcription is complex compared to the simple initiation of transcription in prokaryotes. Enhancers are common in eukaryotes but not in prokaryotes. Transcription and translation occur simultaneously in prokaryotes and separately in eukaryotes. Regulation by siRNAs and miRNAs is absent in prokaryotes but common in eukaryotes.

Explain the role of chromatin packaging in gene regulation and how modifications to chromatin affect gene expression.

-This process determines the accessibility of DNA to transcriptional machinery

-Chromatin remodeling complexes:

Repositioning nucleosomes will allow or limit transcriptional machinery access to DNA

This is targeted to specific DNA sequences by certain TFs

-Histone modification: methylation, acetylation

-DNA methylation

Histone Acetylation

stimulates transcription

Histone Methylation

activates or represses transcription

DNA Methylation

Usually generates 5-methylcytosine near transcription start site (TSS)

Usually represses transcription - may be long term

Summarize the components involved in transcriptional control in eukaryotes and how each contributes to determining whether transcription will occur.

Core promoter and general TFs are necessary but not sufficient

Regulatory promoter/enhancer bind specific TFs -> activators and repressor

Repressors don’t directly block RNA pol instead they compete with activators for DNA site, prevent activators from binding to basal transcription site, bind to silencers

Silencer -> inhibitory version of enhancer

Insulator - DNA sequence that blocks enhancer

Regulation for stalling and elongation -> in some genes RNA pol transcribes 24-50 nucleotides then stalls and transcription will resume in response to a signal, may occur at 30-50% of genes in some eukaryotic genomes

Explain why DNA replication is termed "semi-conservative".

Semi-conservative => each strand can serve as a template

Each containing one parent strand and one new strand

Explain the bidirectional nature of replication

Two replication forks are formed at each ori and move in opposite directions

They first open the double stranded DNA and the DNA template strands are ready for synthesis

Distinguish between the leading and lagging strands.

Leading strand: continuous 5’ -> 3’ synthesis

Lagging strand: discontinuous 5’ -> 3’ synthesis

Describe how replication occurs simultaneously for both strands

During initiation 2 replication forks are formed and move in opposite directions

Primase

makes primer for leading strand and starting point for DNA pol to begin synthesizing new strand

DNA Poly III

elongates DNA

DNA Poly I

removes and replaces primers

DNA Ligase

seals nick and joins new okazaki fragments of lagging strand

Single Stranded DNA Binding Protein (SSB)

maintains lagging strand template DNA as single strand

DNA Helicase

unwinds DNA

Sliding Clamp

keeps DNA polymerase on track

DNA Gyrase/Topoisomerase

induces double strand breaks to prevent torsional strain on DNA

Identify biological processes that involve DNA recombination.

Recombination = the exchange of genetic info between DNA molecules

Allelic shuffling during crossing over in meiosis → increases genetic variation

Bacterial conjugation

Some types of DNA repair

Generation of genetic maps by scientists

Mutation

an inherited change to the DNA sequence and is the source of all new genetic variation. they have the potential to change the expression or function of the encoded protein that causes a change in the phenotype. 

Somatic Mutation

occurs in a non-reproductive cell and is not passed onto the next generation

Germ-Line Mutation

mutation in a sex cell that can be inherited and passed onto the next generation

Base Substitution

one base is converted to another

Transition

purine replaced by purine or vice versa for pyrimidine

Transversion

purine replaced by pyrimidine or vice versa

Insertion

-addition or one or more bases; can lead to frameshift mutation

-example is expanding nucleotide repeats where the repeat is often trinucleotide CNG which may alter gene expression as a result of hairpin structure formed

Deletion

removal of one or more bases; can lead to frameshift mutation

Missense Mutation

a new codon encodes for a different amino acid; there is a change in amino acid sequence

Nonsense Mutation

new codon is a stop codon; there is premature termination of translation

Silent Mutation

the new codon encodes the same amino acid; there is no change in amino acid sequence

Suppressor Mutation

occurs at a site different from that of the original mutation and produces an individual that has both the original mutation and the suppressor mutation but has the wild-type phenotype

Distinguish between incorporated errors and replication errors

an incorporated error is a DNA replication mistake that has been corrected by the cell's proofreading mechanisms, while a replication error is a mistake that has not been corrected and becomes a permanent mutation in the DNA sequence.

Transposable Elements

genes that are able change their position within a genome, nicknamed “jumping genes”. their typical structure contains a terminal inverted repeat with a flanking direct repeat on both ends.

DNA Transposase (class II)

movable component is DNA (DNA is moved and inserted). Encodes for transposase gene and is done through DNA, present in pro and euk

Retrotransposons (class I)

transpose through an RNA intermediate where DNA is copied to RNA which is then reverse transcribed into DNA, needs to have reverse transcriptase gene, present in euk only

What is the mechanism for how transposable elements are able to move around in the genome?

mechanism includes staggered breaks which are generated by transposase, transposon is joined to single stranded ends of target DNA, then DNA in single strand gaps is replicated/filled in-FLANKING DIRECT REPEATS ARE GENERATED DURING INSERTION as these repeats are recognized by enzymes that catalyze transposition

What are the different types of chromosomal variations and how they come about?

duplication, deletion, inversion, and translocation, all of which originate from double-stranded breaks and incorrect rejoining of ends or errors in crossing over.

Duplication

tandem (adjacent), displaced (distant on same or on diff chromosome), or reverse (inverted in direction). this increases gene copy number and can change the phenotype allowing for divergence in gene function and thus evolution.

Deletion

loss of chromosome segments which are apparent as shortened mitotic chromosomes or in paired homologs in meiosis. loss of function is often lethal, but there are low gene dosage effects (haploinsufficiency), loss of dominant allele leading to expression of remaining recessive allele (pseudodominance), or loss of centromere region which leads to chromosome loss.

Inversions

conservation of genetic material is maintained, but sequence is changed, can lead to gene breaks (loss of function) or position effects (i.e. going from an easily transcribable place to a low level place on the chromosome). may be paracentric including centromere or pericentric not including centromere.

Translocations

caused by double strand breaks and incorrect recombination of fragments. An exchange between nonhomologous chromosomes occurs and may be reciprocal (both chromosomes get new DNA) or nonreciprocal (only one DNA gets translocated genes). leads to loss-of-function and creates position effects.

Explain how mitosis allows for the separation of sister chromatids.

First, the nuclear membrane is present and the chromosomes are relaxed. The chromosomes then begin to condense- each chromosome possesses two chromatids. The mitotic spindle forms outside of the nucleus. The nuclear membrane then disintegrates, spindle microtubules attach to chromatids. The chromosomes line up on the metaphase plate. Sister chromatids separate and move toward opposite poles. Chromosomes arrive at spindle poles, the nuclear membrane re-forms and the chromosomes relax.

Explain how meiosis gives rise to haploid gametes.

Meiosis consists of meiosis 1 where the chromosome number is reduced by half and then meiosis 2 where the sister chromatids are separated to produce four separate haploid gametes (equational division). It is important to note that replication occurs prior to meiosis 1 resulting in homologous chromosomes with 2 sister chromatids each. The sister chromatids are no longer identical because of crossing over in prophase 1 (allelic shuffling).

Explain how meiosis increases genetic variation.

Crossing over of homologous chromosomes and random separation of homologous chromosomes.

Gene

an inherited factor (encoded in the DNA) that helps determine a characteristic

Allele

one of two or more alternative forms of a gene

Locus

a specific place on the chromosome occupied by an allele

Genotype

a set of alleles possessed by an individual organism