BIO 340 Exam 4
matastasis
the spread of cancer cells from one part of the body to another - invade normal tissues and travel to new locations, forming new tumors;
mRNA in trpL
the termination stem loop, which halts RNA polymerase progress along the DNA within the leader region
paralogs
homologous genes in the same species
Major predictions of operon model
- lacL gene produces diffusible product (trans-acting) - lacO region involved in regulation - lacO gene region must be adjacent to structural genes to regulate transcription
histone acetyltransferase (HAT)
- recruited by activators and add acetyl(COCH3) groups - These add acetyl groups to positively charged residues in the N-terminal histone tails; the acetylation neutralizes the positive charge and relaxes the histone/DNA interaction - Acetyl groups are removed by histone deacetylases (HDACs), which are recruited by repressors
attenuation of trp operon
- second mechanism controlling trp operon by trpL region
Bases are forced apart to accommodate intercalating agents:
- Ethidium bromide - Acridine orange - Actinomycin D - SYBR® Safe
GAL genes are inducible:
- Transcription is regulated by presence or absence of substrate (galactose) - Absence of galactose: GAL structural genes not transcribed - Galactose added to medium: Transcription begins immediately
autoinducer function
- binds to a regulatory molecule, which in the case of A. fischeri is LuxR. - The LuxR-autoinducer complex then activates transcription of the luciferase target genes that confer bioluminescence.
normal cells
- highly specialized - confined to boundaries of tissue type and contact inhibition
additional control of the lac operon
- inducer: repressor complex alone yields only basal transcription with only a small number of polycistronic mRNAs - Basal transcription is not sufficient to generate enough copies of the lac operon mRNA for metabolism of lactose - Positive control of the lac operon occurs at the CAP-cAMP binding region of the lac promoter - The site attracts the CAP-cAMP complex composed of the catabolic activator protein (CAP) and cyclic adenosine monophosphate (cAMP)
GAL 1 and GAL 10 genes
- transcription of both genes are positively regulated - controlled by central control region UASg (upstream activating sequence of GAL genes) - contain four binding sites for Gal4 protein (Gal4p)
Transcription Control
- under control of a single regulatory region 1. Genes coding for enzymes with regulatory functions are organized in clusters 2. Regulatory regions usually located upstream of gene cluster they control 3. Cis-acting site / domain - regulatory region on same strand 4. Trans-acting elements - molecules that bind cis-acting sites
cAMP (cyclic adenosine monophosphate)
-To bind to promoter, CAP must be bound to cyclic adenosine monophosphate (cAMP) -Glycolysis inhibits activity of adenylyl cyclase, which catalyzes conversion of ATP to little cAMP -when glucose is present, almost no CAP-cAMP can form
Cancer cells share two fundamental properties
-abnormal cell growth and division: unregulated cell proliferation -metastatic spread
cancer cells
-dedifferentiated (look and behave more like primordial cells) - higher rate of proliferation than normal cells - larger with larger nuclei than normal cells - poor organization as they grow
bacterial innate immunity
-express endonucleases (restriction enzymes) which cleave specific dna sequences -restriction enzymes destroy foreign bacteriophage dna, while the bacterium protects its own dna by methylating it -block phage absorption -block phage dna insertion -induce suicide in infected cells
modulation
-mRNA translation - protein processing - modification - degradation
repressible operons in anabolic pathways
-operate through activity of the end product to block transcription of the operon - Certain repressible operons have a second regulatory capability called attenuation which fine-tune transcription to match the immediate needs of the cell
SWR1 complex
-responsible for replacing common histone H2A protein with a variant form (H2AZ)
double helix distortion
-very dynamic structure, flexible
natural circular plasmids range in size from _______. They behave as additional genetic units which are inherited and ________ of the bacterial chromosome, but rely on the _____ and ____ for transcription and replication
1 to > 200 kb replicate independently host enzymes proteins
apoptosis steps:
1. Fragmentation of nuclear envelope 2. Disruption of internal cellular structures 3. Dissolution of cell into small, spherical apoptotic bodies 4. Engulfing of apoptotic bodies by phagocytic cells
selection of transformants
1. LacZ gene codes for the enzyme called β-galactosidase 2. β-galactosidase metabolizes lactose producing galactose and glucose. 3. β-galactosidase converts X-Gal into a blue colored product 4. To activate LacZ gene transcription the activator called IPTG is required
Lac Operon Function and Mechanism:
1. Lactose permease allows the entry of lactose into the E. coli cell from a growth medium. 2. Lactose has its galactoside linkage cleaved by beta galactosidase. 3. Beta galactosidase converts lactose to allolactose. 4. Excess allolactose is cleaved by beta galactosidase into galactose and glucose.
lac (lactose) operon have three features:
1. Regulator 2. Control locus 3. Structural locus
The chromatin modifications alter the strength of the nucleosome-DNA associations, and can cause chromatin structure to either:
1. Relax: forming open promoters 2. condense: leading to closed promoter structures
3 mechanisms which trans-acting proteins access target DNA sequences
1. Some regulatory sequences are not tightly bound by histones, so that regulatory proteins can easily access the DNA sequences 2. Change the distribution or composition of histones by chromatin remodelers 3. Enzymatic modification of histones by addition or removal of acetyl or methyl groups to particular lysine residues
open promoter function
1. The poly A/T tract of open promoters contains binding sequences (BS) that attract transcriptional activators (ACT) 2. The binding region is flanked by sequences that help position one nucleosome upstream and one downstream (called the +1 nucleosome) of the NDR 3. The downstream nucleosome contains a variant histone H2A, called H2AZ, that is easily removed from the transcription start site
Adenomatous polyposis coli (APC)
A tumor suppressor gene. If mutated, it no longer has function. - common in colorectal (colon) cancer.
polymerase chain reaction (PCR) steps
1. Thermostable DNA polymerase 2. Template 3. Primers (short oligonucleotides 18-40 nts) 4. Nucleotides (dNTPs) 5. Buffer (with mono and divalent cations)
low levels of tryptophan
1. Translation stalls at codons 10 and 11 2. The ribosome obscures region 1, allowing regions 2 and 3 to pair 3. Antitermination conformation continued transcription of genes needed for production of tryptophan
when galactose is present:
1. galactose and Gal3 bind to Gal80 2. Gal80 releases Gal4, freeing the DNA-binding domain of Gal4 to recognize and bind to the UASG sites 3. Transcriptional activation domain activates transcription of GAL genes
requirements for vectors:
1. origin of replication 2. selectable marker 3. multicoloning site / polylinker: site that allows gene to be cloned to be placed into the plasmid vector
trp operon contains:
1. regulatory region with promoter (trpP) 2. operator (trpO) 3. leader region (trpL) containing attenuator region 4. structural genes for tryptophan synthesis: trpE, D, C, B, A 5. Gene (trpR) outside operon that encodes repressor protein (trpR) that is activated when bound to tryptophan
enhancers
A DNA sequence that recognizes certain transcription factors that can stimulate transcription of nearby genes. Located on either side of gene, some distance from gene, or even within gene - Important in reaching maximum level of transcription
Similarity searches
A genome sequence statistically similar to gene with known function likely encodes for protein with similar function
metastatic spread
A process by which cells detach from the primary tumor and move to other sites in the body, forming new malignant tumors The ability to invade new tissues results from new mutations in cancer cells
repressors
A protein that suppresses the transcription of a gene, silence element, silence transcription initiation
pair end sequencing
A sequence is generated from both ends of DNA fragments of known size
MicroRNA
A small, noncoding RNA molecule, typically about 21-25 nucleotides long, that binds to mRNA to inhibit its translation. - transcribed by RNA polymerase II and produce transcripts that foldback upon themselves to produce dsRNA -Precursor transcripts are synthesized in the nucleus and processed into miRNAs by Dicer activity
mutations in ras genes lead to production of _____ and ______
hyperactive ras protein, increased cell division
trans-activating domain
Activates or represses transcription by binding to other transcription factors or RNA polymerase
positive control
Activator protein must be present to turn on gene transcription
Human Genome Project (HGP)
An international collaborative effort to map and sequence the DNA of the entire human genome. - clone by clone approach
omics
Areas of biological research having an "omics" connection are continually developing Proteomics Metabolomics Glycomics Toxicogenomics, etc
Enhancer and silencer sequences
At greater distances from the core promoter are the enhancer and silencer sequences (or enhancers and silencers)
Inducible enzymes
Bacteria adapt to environment by producing inducible enzymes only when specific substrates are present
DNA binding domain
Binds to specific DNA sequences in the cis-acting regulatory site
RISC
Binds to the 21-25 bp andseparates the strands to create miRNAs
DNase I hypersensitivity
Hypersensitivity occurs in the immediate vicinity of transcribed genes and surrounding promoters, enhancers, and other transcription-regulating sequences
sporadic cancer
Cancer caused by accumulation of a number of mutations in somatic cells; 90-95% associated Mutation occurs in a single somatic cell
How can a virus cause cancer?
Carry an oncogene into cell and insert it into host genome Altered cell regulation due to changes in kinase activity or production of regulatory proteins Insertion of promoter or enhancer next to cellular oncogene
metastasis enters blood or lymphatic system, and is controlled by large number of gene products:
Cell-adhesion molecules, cytoskeleton regulators, and proteolytic enzymes
chemical
Chilling cells in the presence of divalent cations such as Ca2+ (in CaCl2) renders the cell walls more permeable to plasmid DNA
DNase I hypersensitive sites
Chromatin region that becomes sensitive to digestion by the enzyme DNase I
GAL gene system in yeast (saccharomyces cerevisiae)
Composed of four structural genes and three regulatory genes Products of structural genes transport galactose into cell for metabolism CHECK slide for photo
repressible operons
Contain genes coding for anabolic enzymes - Several genes in a series are turned off (repressed) by the product synthesized by the enzyme
ISWI complex
Contains remodelers that mainly control placement of nucleosomes into a position that silences transcription
Mechanisms of posttranscriptional gene regulation
Control of alternative splicing mRNA stability Translation RNA silencing
operon
Coordinated set of genes regulated as a single unit - Found only in bacteria and archaea - cap site, lac promoter, operator, lacZ, Y, A, lac terminator
Two types of regulatory proteins are involved in cell cycle control are what?
Cyclins Cyclin-dependent kinases (Cdks)
transcription regulatory proteins
DNA binding domain trans-activating domain
epigenetic modifications
DNA methylation Histone acetylation and phosphorylation
CRISPR- Cas
Defenses are part of "adaptive" immunity if the defense mechanism uses past exposure to a pathogen to stimulate improved defense against future exposure to the same pathogen
non specific interactions
Deoxyribonuclease I (Dnase I) binds electrostatically to DNA at the minor grove
computer automated DNA sequencers
Designed for high-throughput sequencing Made genomics possible Essential for Human Genome Project Sequences contained multiple capillary gels Generated over 2 million bp per day
Tryptophan is absent from the cell:
the repressor protein cannot bind the operator thus allowing for transcription of the operon
transformation: eukaryotes
Electroporation Transfection Microinjection Biolistic (gene gun) Ti plasmid - tumor-inducing plasmid of Agrobacterium radiobacter
lacZ (b-galactosidase)
Enzyme that converts disaccharide lactose to monosaccharides glucose and galactose - Conversion is necessary for lactose to serve as primary energy source in glycolysis
LacY (Lactose Permease)
Enzyme that facilitates entry (PERmits) of lactose into bacterial cell
lacA (galactoside transacetylase)
Enzyme that may be involved in removal of by-products of lactose digestion from the cell
constitutive enzyme
Enzymes are continuously produced regardless of chemical makeup of environment
qPCR- TaqMan probe
Fluorescence-labeled oligonucleotides - 5' to 3' exonuclease activity of the polymerase cleaves the probe, releasing the fluorophore into solution, where it is allowed to fluoresce
whole-genome shotgun sequencing (WGS)
Focuses on sequencing all of the DNA in an organism's genome by cutting the DNA into many small, easily sequenced sections then uses computer algorithms to align overlapping segments to assemble the entire genome.
Phases of the cell cycle
G1, S, G2, M Three important checkpoints: G1, G2, and M phases - G1 checkpoint is most important
orthologs
Genes from different species thought to have descended from common ancestor
oncogenes
Genes that induce or continue uncontrolled cell proliferation
proto-oncogenes
Genes that initiate or maintain cell division May become cancer genes (oncogenes) bymutation
IPTG - isopropylthiogalactoside
Gratuitous inducers are analogs of lactose
familial retinoblastoma
Individuals inherit one mutant copy of RB1 gene • 85% to 95% chance of developing the disease
inherited cancer
Inherited genes cause a predisposition to cancer; 5% associated with germ line mutations Mutations are carried in all cells
coactivators
Interact with proteins and enable activators to make contact with promoter-bound factors ; form complex enhanceosome
RNAi is widespread in eukaryotes
It is thought that it evolved by helping organisms protect their genomes against mutational effects of transposable genetic elements
recombinant DNA
Joining of DNA molecules - Produced by artificially joining DNA from different biological sources not found together in nature
absence of lactose
LacI binds as a tetramer to the operator region. It represses the lac operon by preventing open complex formation by RNA polymerase. NO lactose = Enzymes are not needed and expression of genes encoding enzymes is repressed, lac operon and transcription is blocked
____ stimulates transcription of lac operon and induces synthesis of enzymes
Lactose
RB1 tumor suppressor gene
Loss or mutation of RB1 gene contributes to development of cancers due to unregulated progression through cell cycle Originally identified via studies on retinoblastoma: rare retinal cancer
chemical modification of chromatin
Lysine is frequently targeted for methylation Methylation plays a role in converting open chromatin to closed chromatin Demethylation is carried out by histone demethylases (HDMTs)
eukaryotic gene regulation
More complex than that in prokaryotes - Large amount of DNA is associated with histones and other proteins - mRNAs must be spliced, capped and polyadenylated prior to transport from nucleus• - Genes on numerous chromosomes
Methyl (CH3) groups are added to _____ by _______
N terminal histone tails, histone methyltransferases (HMTs)
inducible operon
Operons encoding enzymes that act in catabolism• - Operon is turned on (induced) by substrate(s) for which the structural genes encode - Enzymes needed to metabolize a nutrient are only present when that nutrient is present in the environment
tumor inducing plasmid (Ti plasmid)
Part of the plasmid —transfer DNA (T-DNA) is transferred from the bacterium to plant cell nuclei upon infection - Once inside the plant cell, T-DNA integrates into random locations of the plant genome
transformation: prokaryotes
Plasmids are introduced into bacteria via transformation and hosts are engineered to lack restriction enzymes and recA
CAP (Catabolite-activating protein)
Positive control of the lac operon occurs at the CAP binding site of the lac promoter - The site attracts the CAP-cAMP complex composed of the catabolic activator protein (CAP) and cyclic AMP (cAMP) - Binds to CAP-binding site, facilitating RNA polymerase binding at promoter and facilitating transcription
polymerase chain reaction (PCR)
Process of increasing small quantities (amplifying) of DNA for analysis - Used for diagnostic tests for genetic diseases and detecting pathogens - Reverse-transcription PCR (RT-PCR) uses mRNA as template
control locus
Promoter: recognized by RNA polymerase Operator: acts as an on/off switch for transcription
caspases
Proteases responsible for initiating apoptosis and digesting intracellular components
readers
Proteins that recognize modified histones
activators
Proteins that turn operons on by binding to DNA; enhancer element, enhance transcription initiation
Sporadic retinoblastoma
RB1 gene mutation on chromosome 13
clones
Recovered copies of recombinant DNA molecule - Used to study structure and orientation of DNA
tumor repressor proteins
Repair damaged DNA Control cell adhesion Act in cell-signaling pathways that inhibit the cell cycle
silencers
Repress the level of transcription initiation
catabolite repression
Without CAP-cAMP bound to the lac promoter region, lac gene transcription is very inefficient
because double helix are flexible aromatic macrocyclic flat _____ composed of fused, heterocyclic rings, can ____ between the _____ pairs of bases
hydrophobic molecules, slip, stacked
trpL
Short polypeptide with two back-to-back codons for tryptophan - "sensors" for the availability of tryptophan in the cell Encodes a short polypeptide called the Leader peptide contains four sequences regions (1,2,3,4) and the mRNA produced can form loop structures
Human genomic sequence is 99.9% same with most genetic differences resulting from:
Single nucleotide polymorphisms (SNPs) Copy number variations (CNVs)
genomic alternations associated with cancer include:
Single-nucleotide substitutions Chromosomal rearrangements Deletions
SWI/SNF complex
Slides and relocates nucleosomes. Also functions to open chromatin structure by displacing or ejecting nucleosomes.
epigenetics
Study of factors that affect gene expression but do not alter nucleotide sequence of DNA
co-suppression
Suppressed expression of both endogenous gene and transgene
tumor-suppressor genes
Suppression of the cell cycle can be important in the case of damage to a cell'sDNA - p53 prevents a cell from passing on mutations due to DNA damage - Mutations in the p53 gene prevent suppression of the cell cycle - p53 is the most frequently mutated gene in cancers (50% of all cancers)
microsynteny
Synteny at the level of just a few genes
core promoter region
The core promoter region containing the TATA box and other sequences. -It is immediately adjacent to the start of transcription -It is the sequence to which RNA polymerase II and its associated transcription factors bind
loops in enhanceosomes
The loops allow enhanceosome proteins to interact with RNAP II and transcription factors at the core promoter and proximal promoter elements Loops may be small or large based on the distance between the enhancer sequence and the gene it regulates
sequence specific interactions
The major groove is more accessible for highly specific interactions
lac repressor function
The three segments of the operator share a conserved 21-bp inverted repeat around a central G-C base pair - The tetrameric repressor binds to O1 and O3 and induces DNA loop formation that brings O1 and O3 close together - The loop structure includes part of the promoter and blocks access by RNA polymerase
proto-oncogenes can be converted to oncogenes by:
Translocation gene duplication point mutations
RNAi can silence gene expression transcriptionally and posttranscriptionally T/F?
True
writers and erasers are recruited to specific chromatin locations by DNA binding proteins to modify histone tails, T/F?
True
enhancer element is called the upstream activator sequence (UAS or UASg) T/F?
True - each UASg contains two 17 bp repeat sequences that are binding sites for Gal4
_____ acts as a correpressor by binding to the trp repressor and activating it
Tryptophan
BLAST searches
Used to screen databases and compare a sequence to a known sequence
Single nucleotide polymorphisms (SNPs)
Variations in the DNA sequence due to the change of a single base in genome
cloning vectors
a DNA molecule that can accept foreign DNA, be transferred to a host cell, and get replicated in it - plasmids - phases and viruses - cosmids - artificial chromosomes
electroporation
a brief electrical pulse applied to a solution containing cells creates temporary pores in their plasma membranes, through which DNA can enter
transformation
a normal cell converted to a cancerous cell
autoinducer
a small signal molecule that takes part in quorum sensing
Taq DNA polymerase
a thermostable polymerase used in polymerase chain reaction isolated from Thermus aquaticus - half life is 40 mins at 95 celsius - Low replication fidelity because it lacks a 3' → 5' exonuclease proofreading mechanism
gene regulation
ability of an organism to control which genes are transcribed in response to the environment/ turning on and off genes
Conversion of a proto-oncogene to anoncogene can lead to ____ of the cell cycle
abnormal stimulation
dysplasia
abnormal/disorganized fast development or growth of cells, tissues, or organs -progress to neoplasia
_____ binds to trpO and prevents transcription of the operon
activated repressor
writer enzymes
add chemical groups to chromatin
Principal chemical modifications are _____ and removal of ______ and ______ groups
addition, acetyl , methyl
restriction endonucleases (restriction enzymes)
an enzyme produced by certain bacteria, having the property of cleaving both strands of DNA molecules at or near a specific sequence of nucleotides.
enhanceosomes
are protein complexes that assemble at enhancers to facilitate transcription. - direct DNA bending into loops
presence of lactose
b-galactosidase (LacZ), when at low levels, cleaves and rearranges lactose to make the inducer allolactose. Allolactose binds to LacI, reducing its affinity to the operator and thus allowing induction of the operon. Lactose present: Indirectly induces activation of genes by binding repressor
negative control
binding of a repressor protein to a regulatory DNA sequence and preventing transcription of a gene or gene cluster
Transcription negative control:
binding of a repressor protein to a regulatory DNA sequence and preventing transcription of a gene or gene cluster - Some repressor proteins undergo inactivation - Some repressors undergo activation
positive control
binding of an activator protein to regulatory DNA and initiating gene transcription
Transcription positive control:
binding of an activator protein to regulatory DNA and initiating gene transcription -Allosteric effector compound binding induces a conformational change that activates the DNA-binding domain -Activator proteins have a DNA-binding domain that is converted to in active conformation by binding of an inhibitor to the allosteric domain
repressor has: Allosteric domain
binds a molecule or protein and cause a change in conformation of the DNA-binding domain - allostery
in plants, infection of a single leaf by a virus can generate RNAI response that _______
blocks viral replication and protects the rest of the plant from the infection
BRCA1 and BRCA2 genes increase susceptibility to _____ and ____ cancer
breast, ovarian - BRCA1 and BRCA2 are DNA damage repair genes
programmed cell death
called apoptosis - occurs when DNA /chromosome damage is too sever to repair so cells stop progressing in the cell cycle - prevents cancer - eliminates cells not contributing to final adult organism
cancer-causing agents
carcinogens; - any substance or event that damages DNA and causes mutationsto occur in proto-oncogenes or tumor-suppressor genes
open promoters
cause genes to be constitutively expressed - they have a nucleosome-depleted region (NDR) of 100-150 bp immediately upstream of the transcription start site
covered promoters
characterize genes whose transcription is regulated -transcription is blocked until nucleosomes are displaced or removed from the promoter - There is active competition between nucleosomes and transcription factors for binding
cloning into plasmids
check slides page 18
carcinogens are:
chemcials (natural of sythetic) radiation some viruses
chromatin modifier proteins
chemically alter histone proteins in the nucleosomes by adding or removing chemical groups
binding of repressor proteins to silencer sequences
cis-acting regulatory sequences that directly prevent enhancer-mediated transcription
regulator
composed of the gene that codes for the repressor (lacI) - a protein capable of repressing the operon
hallmarks of cancer
concepts of proto-oncogenes and tumor suppressor genes alone cannot fully describe genetic abnormalities driving cancer development and progression
Drosha enzyme complex
cuts the pri-miRNA near the middle of the stem and produces two segments
DNA ladders or markers
designed for sizing of double-stranded DNA made by restriction digestion of DNA with restriction enzymes
restriction map
diagram that shows the lengths of fragments between restriction sites in the strand of DNA - Provides information on length of cloned insert and location of restriction sites within clone - smallest fragments move farthest down the gel - fragments visualized on gel
fluorescence is _______ proportional to the amount of template at that time
directly
ras genes
encode signal transduction molecules that regulate cell growth and division in cell membrane; most frequently mutated genes in human tumors
activator protein binds at
enhancer element that favor formation of PIC and initiation of translation
transcription of genes in the galactose utilization pathway is carefully regulated by _____
enhancer-like sequences
Gene Silencing by double stranded RNA: Dicer
enzymes that cuts the dsRNA into 21-25 bp fragments of siRNA or miRNA that are bound by RNA-induced silencing complex(RISC), which denatures them
attenuation
fine-tune transcription to match the immediate needs of the cell
SYBR safe
fluorescent dye that is positively-charged and binds non-specifically to DNA predominantly via electrostatic interactions and groove binding
blunt ends:
fragments produced with double-stranded ends
Sticky ends (cohesive ends)
fragments produced with overhangs
GTFs
general transcription factors; trans-acting regulatory proteins, which can bind to their target sequences on any chromosome
homologous genes
genes that are evolutionarily related
the protein Mig1 is produced in the presence of ____ and binds ______ upstream of GAL genes
glucose, silencer sequences
tumor suppressor genes
help prevent uncontrolled cell growth
Most transposons in eukaryotes are located in ____
heterochromatin and are silent
Eukaryotic repressors _____ through different mechanisms than those seen in bacteria
inhibit transcription
pri-miRNA
initial transcript is called primary microRNA; forms double stranded stem of 65-70 nucleotides
defining feature of eukaryotic DNA is _____
its packing into chromatin
proximal elements
located near and upstream of the promoter, regulates genes
repressor has : DNA-binding domain
locates and binds operator or other target DNA sequences
structural locus
made up of three genes, each coding for a different enzyme needed to catabolize lactose 1. lacZ: b-galactosidase 2. lacY: lactose permease 3. lacA: galactoside transacetylase
divalent cations
magnesium or manganese ions - The divalent ions are co-factors of Taq DNA polymerase activity
alternative splicing
many genes code for multiple proteins - HGP found that number of genes is lower than number of predicted proteins
malignant tumors
metastasized cells invading other tissue and causing life-threatening problems
chromatin remodeling
modifications that reposition nucleosomes so as to open or close promoters and other regulatory sequences
codon bias
multiple codons for the same amino acid are not used equally; nonrandom - due to scarcity of some tRNAs - Organisms vary in the degree to which certain codons are used when there is more than one codon for a given amino acid
RISC (RNA-induced silencing complex)
multiprotein complex that contains aprotein in the Argonaute family
results from mutated gene products or abnormally expressed genes:
mutations continue to accumulate; mutations affect multiple cellular functions
delay between exposure to carcinogen and appearance of cancer is an indication of the _____
mutlistep process - ex: leukemia from radiation exposure has incubation period of 5-8 years
DNA and RNA is ____ charged DNA can be _____ and form using gel electrophoresis
negatively separated based on size
cancerous cell is characterized by at least one active _____ and mutation of _____
oncogene , tumor suppressor genes
dideoxynucleotide chain termination sequencing (sanger)
one strand of a DNA fragment is used as a template for synthesis of a nested set of complementary fragments
transgenic plants
plants are infected with bacteria whose plasmids usually cause tumors in plants. the tumor producing gene is inactivated and the bacterial plasmid instead only introduces the desired donor gene - The opine- and tumor-producing genes of T-DNA can be replaced by a gene of interest - The gene is inserted between the left and right borders of the T-DNA
Posttranscriptional regulation
plays an important role in regulating gene expression via alternative splicing
Regulation of inducible or repressible type transcription is under _____ or _____
positive control or negative control
monovalent cations
potassium, sodium and ammonium ions - Monovalent ions stimulate the activity of Taq DNA polymerase and shield the negatively charged phosphate backbone, thereby weakening the electro-repulsive forces between primer and target DNA
closed chromatin
produced by modifications that cause regulatory sites to be covered by nucleosomes, restricting access by regulatory proteins
Encyclopedia of DNA Elements (ENCODE)
project seeking to determine transcriptional start sites, promoters, enhancers, and functional elements in the human genome
Gal4
promoter and enhancer elements bind to regulatory activator protein Gal4 - continuously present in cells - interacts with Gal80 protein and keeps it inactive when galactose is absent
Burkitt's lymphoma
reciprocal translocation between chromo 8 and 2, 14, or 22 -over expression of MYC protein drives excessive cell division
Chronic Myelogenous Leukemia (CML)
reciprocal translocation between chromo 9 and 22 - both mature and immature granulocytes are overproduced in the marrow and blood
open chromatin
relaxed association between DNA and nucleosomes and allows for access by regulatory proteins
eraser enzymes
remove groups from chromatin
high tryptophan levels
ribosome moves to stop codon and region 1 binds to region 2, which leads to the binding of region 3 and region 4, causing termination of transcription prematurely by attenuation
Paradigm of the lactose operon
scientists Jacques Monod and François Jacob proposed the idea that genes could be regulated in E. coli, enzymes used to metabolize - Lactose were inducible - Glucose were constitutive -lactose utilization lacZYA operon of E. coli was the first gene regulatory system described
Copy number variations (CNVs)
segments of DNA that are duplicated or deleted
repressor proteins bind at the ________
silencer elements at a decrease rate of preinitiation complex (PIC) assembly and RNA II polymerase release
plasmids
small circular extrachromosomal double stranded DNA molecules that replicate independently from the bacterial chromosome
promoters
specific region of a gene where RNA polymerase can bind and begin transcription made up of DNA sequence elements: - initiator (Inr) - TATA box -TFIB recognition element (BRE) -Downstream promoter element (DPE) -Motif ten elment (MTE)
chromatin remodeling and modifying enzymes are recruited to
specific sites by trans-acting factors that bind target DNA sequences
SSU rRNA gene
standard method for identifying species
Metatranscriptomics
study of RNA transcripts obtained from an environmental community
functional genomics
study of gene function based on the mRNAs or possible proteins they encode as well as regulatory elements
Metaproteomics
study of protein data obtained from an environmental sample
metagenome
sum total of all DNA sequenced from an environmental sample
antibiotic selection
the process of culturing bacteria on antibiotic-containing medium, which only allows transformed bacteria to grow - To select recombinants / clones, most plasmids havean antibiotic resistance gene
quorum sensing
the process where bacterial cells work together at high density - discovered in Aliivibrio fischeri, a bioluminescent bacterium
multiple _____ factors bind to several different enhancer and promoter elements and fine tune the level of transcription initiation
transcription factors
cis-acting sequences
transcription regulation requires binding of many regulatory factors to specific DNA sequences: promoters, enhancers, silencers - located on same chromosome as gene that it regulates
pre-miRNA
transported to the cytoplasm where dicer removes the terminal loop
____ strains have ___ rates of operon transcription in the presence of tryptophan than trpR+ suggesting a second mechanism for trp operon regulation
trpR-, higher
trpR- cells should produce ______ constitutively as there is no repressor protein to inhibit transcription
tryptophan
retinoblastoma protein (pRB)
tumor-suppressor protein that controls the G1/S cell-cycle checkpoint - prevents passage into S phase - depends on phosphorylation state
Cancer cells not eliminated by the immune system form ____
tumors
synteny
two or more genes are located on the same chromosome and are physically linked
benign tumors
unregulated cell growth that forms a multicellular mass, removed by surgery, no serious harm
RNAi plays protective role in response to _____
viral infection