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What is the Promoter?
The site where the RNA polymerase binds the DNA to start transcription
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What is the TATA box?
- Binding site of a general transcription factor: TBP or TATA-binding protein
- 25-30 nucleotides upstream of the transcription start sites in eukaryotes
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What are the transcription factors for eukaryote gene regulation?
Proteins that facilitate transcription when bound to the RNA polymerase
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What is required for transcription of all proteins?
General transcription factors
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What are other common promoter elements in eukaryotes?
- CAAT (cat) box, -75
- GGGCGG box (GC box), -90
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What are Activators?
proteins that bind to enhancers and stimulate transcription
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What are Enhancers?
- Regulatory DNA sequences that may be thousands of nucleotides upstream of the gene they regulate
- A gene can have multiple enhancers, each activate at a different time in different tissues
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What are Repressors?
Proteins that Inhibit transcription of a gene
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How do Repressors Inhibit Transcription?
- Blocking activator binding to enhancers
- Binding control elements in an enhancer to turn of transcription even with an activator protein present
- By influencing chromatin structure, for example by recruiting histone acetylase at a site
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Describe the model of transcription initiation in eukaryotes?
- Transcriptional activators bind to DNA and recruit chromatin remodeling complexes and histone acetyl transferases (HATs)
- Chromatin remodeling complexes and histone HATs open the chromatin and expose the regulatory sequences: promoters and enhancers.
- Activator protein bind to enhancer elements, basal transcription factors bind to the promoter and recruit RNA polymerase II
- Mediator proteins connect the activators and basal transcription factors. RNA pol II begins transcription
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What determines the rate of transcription?
- Transcription factors
- Activators
- Repressors
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What is RNA processing?
Splicing and RNA editing post-transcriptional regulation
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What is RNA interference (RNAi): mRNA degradation and blocking translation post - transcriptional regulation
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Describe the Regulation of gene expression by microRNA's (miRNAs).
- The micro-RNA (miRNA) precursor folds back on itself, held together by hydrogen bonds
- An enzyme called Dicer moves along the double-stranded RNA, cutting it into shorter segments
- One strand of each short double-stranded RNA is degraded; the other strand (miRNA) then associates with a complex of proteins
- The bound miRNA can base-pair with any target mRNA that contains the complementary sequence
- The miRNA-Proteins complex prevents gene expression either by degrading the target mRNA or by blocking its translation.
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Describe Degradation of a protein by a proteasome in Eukaryote gene regulation.
- Multiple ubiquitin molecules are attached to a protein by enzymes in the cytosol
- The Ubiquitin-tagged protein is recognized by a proteasome, which unfolds the proteins and sequesters it within a central cavity
- Enzymatic components of proteasome cut the protein into small peptides, which can be further degraded by other enzymes in the cytosol
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What is Ubiquitin?
- A small (76 amino acids) very conserved proteins found in eukaryotic cells
- Main function is to target other proteins for destruction
- Ubiquitin marked proteins are found by proteasomes:
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What Mutations cause cancer?
- Spontaneous point mutations
- Mutations caused by exposure to carcinogens; chemicals that induce mutations
- X-rays, UV light, ionizing radiation
- Viruses that insert themselves into the genome
- Genomic rearrangements (Inversion, translocation, deletion, duplication and gene amplification)
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What is an Oncogene?
A gene that has the potential to turn a normal cell into a tumor cell
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What is a Proto-oncogene?
- Mutations involving these genes can turn them into oncogenes
- Normal genes that stimulate cell division when expressed
- Mutations that lead to excessive expression of proto-oncogenes can turn them into oncogenes.
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What is RAS?
A G protein (GTP binding regulatory protein) and is active when bound to GTP)
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What are Primers?
Primers are short single stranded DNA molecules that are complementary to the ends of the sequence to be amplified. The primers define the what sequence will be amplified and they are also required to initiate replication
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How is a Primer extended?
Thermostable DNA polymerase replicates the template DNA. The DNA polymerase is derived from the thermophilic bacteria Thermus aquatics and is generally referred to as Taq polymerase
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Who earned the Nobel in Chemistry in 1993 for the polymerase chain reaction?
Kary Mullins
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What is the process of whole-genome shotgun approach to sequencing?
- Cut the DNA from many copies of an entire chromosome into overlapping fragments short enough for sequencing
- Clone the fragments in plasmid or phage vectors
- Sequence each fragment
- Order the sequences into one overall sequence with computer software
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What did Barbara McClintock receive a Nobel prize for?
For her results that suggested to her that some genetic factors were capable of changing locations on the chromosome
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What are Insertion Sequences (IS elements)?
- The simplest transposable elements in prokaryotes
- Carry a single that codes for an enzyme (Transposase) That catalyzes their transposition
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What is Transposition?
Movement of genetic elements to new locations
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What are inverted repeats?
- Flanking the transposes gene in Insertion Sequences are inverted repeats
- The Nucleotide sequence of an inverted repeat on one side of the transposes gene is inverted relative to the repeat on the other side of the transposes gene
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How do IS elements generate mutation?
- Inserting ino genes
- Inserting into regulatory regions of a gene and changing the expression of the gene
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What group of DNA does Transposable element belong?
- Selfish DNA
- Only reason for existence is to make copies of self
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What is Duplicative (Replicative) Transposition?
The transposable elements makes a copy of itself when it moves to a new location. Copy and past
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What is Non-replicative (Conservative) Replication?
The transposable element does not make a copy of itself when it moves to a new location. Cut and past
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What are some examples of High throughput (NextGen) DNA sequencing?
- 454 Pyrosequencing
- Illumina/solexa
- Ion Torrent
- SOLiD
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What is Chromatin?
The DNA-Protein complex found in eukaryotes
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What is a tumor suppressor?
A gene associated with tumor formation when its product does not function
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In Eukaryotes, why are certain genes expressed only in certain types of cells?
Different cell types have different regulatory transcription factors
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What is the alternative splicing?
mRNA processing events that lead to different combinations of eons being spliced together
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What type of proteins bind to promoter-proximal elements?
Regulatory Transcription Factors
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Imagine that you have isolated a yeast mutant that contains a constitutively (Constantly) active histone deacetylase. What phenotype do you predict for this mutant?
The mutant will show low levels of gene expression
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The reason for difference in the sets of proteins expressed in nerve cells and pancreatic cells of the same individual is that nerve cells and pancreatic cells contain different ___?
Sets of regulatory proteins
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IN the roundworm C. Elegans, the kin-4 gene produce an RNA that forms a hairpin structure. One of the strands in the double stranded region of the kin-4 hairpin is complementary to the mRNA of protein coding gene , pin-14. Predict the effect of expressing kin-4 RNA during development?
The lin -14 expression will fall when lin 4 expression begins
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In a paper on a gene called BDNF (Brain-derived neurotrophic factor) the authors state that "BDNF is encoded by a complex gene with four well-characterized promoters that give rise to at least eight different mRNA's." What mechanism could account for the production of these different BDNF mRNAs?
Alternative splicing
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_______ bind to enhancer regions?
Activators
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If DNA were inflexible, which of the following would not function?
Enhancers
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If the sequence of a cDNA matches a DNA sequence in the genome, then this genomic DNA is likely to?
Code for protein
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What is the difference between an open reading frame (ORGF) and a gene?
An ORF is a potential gene identified by a potential protein-coding segment in DNA; a gene is a DNA segment known to produce a product
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What mutation converts a proto-oncogene into an oncogene?
A mutation that greatly increase the amount of proto-oncogene protein
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What DNA sequence would qualify as a terminal repeats of bacterial IS element?
It has to be complimentary to each other
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Bacteria containing a plasmid that has the eukaryotic gene integrated into its bgIII site would grow in ____?
The kanamycin broth and the nutrient broth
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The bacteria that did not take up any plasmids would grow on which media?
The nutrient broth
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Bacteria with the plasmid, but without the eukaryotic gene , would grow____?
In all four types of broth
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A major obstacle to expressing (Correctly transcribing and/or translating) unmodified vertebrate genes in bacteria is that______?
Bacteria don't have spliceosomes and cannot remove eukaryotic introns
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Approximately what portion of the DNA in the human genome codes for proteins or functional RNA?
2%
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If alternative splicing did not take place, we would expect that?
The human genome would contain more genes
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Gene expression is often assayed by measuring the lvl of a particular mRNA produced from a particular gene. If one is interested ion knowing the amount the amount of a final active gene product, a potential problem of this method of assessing the level of gene expression is that it is blind to the possibility of?
Translational control
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A functional p53 activates genes that?
Arrest the cell cycle and increase DNA repair
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Which of the following is critical to production of human growth hormone in E. coli?
Th genetic code is universal
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Restriction enzymes are to bacteria what ____ is(are) to mammals
An immune system
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Genetic engineering techniques have developed by borrowing knowledge from unexpected sources. One such source was studies of retrovirus replication. What aspect of a common cloning procedure makes direct use of knowledge?
Synthesizing cDNA's
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What DNA sequence information is critical to the success of PCR?
The sequence of the ends of the DNA to be amplified must be known
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If one wished to test the hypothesis that humans and chimps differ due to differences in the expression of a set of shared genes, the technique to use would be?
Transcriptome sequencing (RNA-Seq)
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What is the leading hypo thesis to explain the paradox that large, complex eukaryotes such as humans have relatively small numbers of genes?
Alternative splicing of mRNA's
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If cells of an individual contain the same set of genes, how do these cells become different from each other during development?
Differences in extracellular signals received by each cell lead to differences in the types of regulatory proteins presents in each cell
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Gene expression is often assayed by measuring the level of a particular mRNA produced from a particular gene. If one is interested in known the amount of a final active gene product, a potential problem of this method of assessing the level of gene expression is that it is blind to the possibility of?
Translational control
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When sequencing genomes by shotgun sequencing, researchers carry out?
DNA sequencing of small fragments and then ordering the sequence of each fragment using overlapping sequencing identity
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In recombinant DNA methods, the term vector can refer to?
A plasmid used to transfer DNA into a living cell
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What double stranded DNA sequence is most likely to be recognized as a cutting site by a restriction enzyme?
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Why do histones bind tightly to DNA?
Histones are positively charges, and DNA is negatively charged
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An important difference between the ways that bacteria and multicellular eukaryotes regulate gene expression is that?
Multicellular eukaryotes need to respond to signals from other cells much more than do bacteria
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Which of the following is most likely to have a small protein called ubiquitin attached to it?
A proteins that needs to be degraded
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Which of the following allows more than one type of protein to be produced from one gene?
Alternative forms of RNA splicing
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The addition of acetyl groups to histones is associated with?
Chromatin decondensation and higher levels of gene expression
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Activator proteins in eukaryotes usually have a domain that binds to DNA and other activation domains that often bind to?
Other regulatory proteins
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An enhancer_____?
Acts at a distance from the promoter
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Differential gene expression is the result of different cells containing different _____?
Regulatory proteins
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The phenomenon in which RNA molecules in a cell are destroyed if they have a sequence complementary to an introduced double-stranded RNA is called?
RNA interference
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The dideoxynucleotides (ddNTPs) used in DNA sequencing work to ____?
Stop the growth of a NDA strand at a particular base
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The DNAs produced in a DNA sequencing reaction are analyzed on the basis of their?
Size or length
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To isolate a cloned gene from a DA library, researchers____?
Probe the library using a labeled single-stranded DNA complementary to the gene
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How do we describe transformation in bacteria?
Assimilation of external DNA into a cell
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What controls which DNA is amplified in PCR?
The choice of primers
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Each round of PCR?
Doubles the amount of DNA
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Why is "chain reaction" an appropriate part of the term PCR?
Newly synthesized DNA segments serve as templates in subsequent cycles
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In gene-cloning projects, plasmids function to ?
Allow replication of the DNA thats being cloned
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A laboratory might use dideoxyribonucleotides to?
Sequence a DNA fragment
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Bioinformatic can be used to scan for short sequences that specify known mRNAs, called ___?
Expressed sequence tags
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Because they both produce a reverse transcriptase, long interspersed nutlear elements as transposable elements may be related to __?
Retroviruses
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In establishing paternity by DNA fingerprinting, what proportion of micro satellite repeat alleles in a child come from the father?
1/2
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A____ is a 2-6 base pair repeated sequence in DNA?
Microsatellite
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To identify protein-coding genes, how must computer programs scan the sequence of a genome?
Scan all three reading frames on each of the DNA strands
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In shotgun sequencing of a complete genome, why is it essential for DNA fragments of overlap?
It allows the sequence date to be put together in the correct order
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A multi gene family is composed of ____?
Genes whose sequences are very similar and that probably arose by duplication
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The 'transciptome' can be defined as?
The complete set of DNA sequences that are transcribed in a particular cell
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Proteomics is defined as the ___?
Study of the full protein set encoded by genome
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What is the central dogma?
DNA makes RNA and RNA makes protein
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Describe the termination of transcription in prokaryotes?
RNA polymerase transcribes through the terminator sequence, causing the polymerase to release the transcript
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A mutation in which of the following parts of a gene is most detrimental to the protein that the gene codes for?
exon
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Which of the following statements are true about proteins synthesis in prokaryotes?
Translation can begin while transcription is still in progress
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What is an Allele?
One of two or more alternative ford of a gene that arise by mutation and are foun at the same place on a chromosome
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A cell divides to produce two genetically identical daughter cells is true when?
Only during mitosis
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Independent assortment of chromosomes occurs when?
Only during meiosis I
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The process is preceded by replication of DNA occurs when?
Both mitosis and meiosis I
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When Mendel crossed true breeding yellow-seeds and green-seeded pea plants what was his expected ratio?
3:1
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When Mandel took the F1 Yellow-seeded plants and crossed them to green-seeded plants, what phenotypic ratio was expected ?
1:1
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There is good evidence for linkage when?
Genes do not segregate independently during meiosis
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