Genetics Lab Final
What do constitutive genes do?
Frequently, constitutive genes encode proteins that are continuously necessary for the survival of the organism
Central dogma refers to the process of what?
Gene expression
forward genetics
Traditional approach to the study of gene function that begins with a phenotype (a mutant organism) and proceeds to a gene that encodes the phenotype. (Phenotype known)
Forward genetics application in lab:
Used multiple generations of C. elegans to determine at what life stage a C. elegan might inherit the trait of a protruding vulva.
What is differentiation?
process in which cells become specialized in structure and function
Parts of a transgene:
- Promoter/enhancer: confers the desired spatial and temporal pattern of transgene expression - 5' and 3' UTR: genomic sequences with strong termination & poly A signals - Introns: increase efficiency of RNA processing and nuclear transport associated with RNA splicing
Benefits of regulating gene expression
1) The benefit of regulating genes is that encoded proteins will be produced only when required 2) Important for metabolism, cell division, and environmental stress responses
Importance of model organisms (particularly C. elegans)
1. Able to reproduce quickly 2. Large % of genome is known 3. Cheap/easily accessible
How does PCR work?
1. Denature DNA using heat (95°C) - separates DNA strand 2. Annealing (55°-65°C) - Adds complementary primers to both the template and complementary strands 3. Extension (72°C) - uses Tag DNA polymerase (heat resistant enzyme) to add/sequence all strands using free nucleotides To amplify a segment of DNA using PCR, the sample is first heated so the DNA denatures, or separates into two pieces of single-stranded DNA. Next, an enzyme called "Taq polymerase" synthesizes - builds - two new strands of DNA, using the original strands as templates
Calculating Chi-square value
1. For each observed number in the table subtract the corresponding expected number (O — E). 2. Square the difference [ (O —E)2 ]. 3. Divide the squares obtained for each cell in the table by the expected number for that cell [ (O - E)2 / E ].
types of transcription factors
1. General transcription factors 2. Regulatory transcription factors
Mechanisms that influence Evolution:
1. Natural Selection 2. Genetic Drift (large population size) 3. Mutations 4. Gene Flow 5. Random Mating If any ONE of these assumptions is not met, then genetic equilibrium is broken and EVOLUTION OCCURS.
blood types
A - has A antigens on the surface of the RBCs and B antibodies (anti-B) in the blood plasma B - has B antigens on the surface of RBCs and A antibodies (anti-A) in the blood plasma AB - has both A and B antigens on the surface of RBCs, but no plasma antibodies. O - has no antigens on the surface of RBCs but has both A and B antibodies in the blood plasma. Type O is the universal donor and AB blood is known as the universal recipient.
Duchenne muscular dystrophy
A human genetic disease, occurring mostly in men/children, caused by a sex-linked recessive allele; characterized by progressive weakening and a loss of muscle tissue due to a mutation in a gene that codes for the protein dystrophin.
Amino acid ratios
Amount of A = Amount of T, Amount of G = Amount of C
Cystic fibrosis
An autosomal recessive genetic disorder; characterized by an excessive secretion of mucus and consequent vulnerability to infection; caused by any one of more than 2,000 mutations in a gene that codes for the CFTR protein..
What do activators do?
Bind to DNA and increase transcription
What do repressors do?
Bind to DNA and inhibit transcription
How does changing the amino acid sequence on your model affect the protein model?
Changing the amino acid changes the way the amino acids interact and changes the shape of the protein.
Explain how changing the shape of the hemoglobin proteins affects the hemoglobin function.
Changing the shape of the protein causes the hemoglobin molecules to stick together to from long rods causing a decrease in blood flow
How will denaturing a protein affect its ability to carry out its function?
Denaturing a protein will change its shape. When this happens the protein may not have the correct shape to carry out its function.
What does an activator bind to?
Enhancer
Exon skipping in DMD
Exon skipping is a technology that changes how the primary RNA transcript of a gene with a disease-causing mutation is spliced, removing the mutation from the resulting mRNA. A mutation that blocks translation results in no protein, or a nonfunctional protein fragment. Exon skipping allows a shortened, but partially functional protein to be produced. (RNA Splicing)
Gene Switches
Gene switches regulate the expression of genes by binding to different proteins that affect the activity of RNA polymerase. These switches bind regulatory proteins that turn the *TRANSCRIPTION* of genes on or off. (Transcription)
Gene therapy
Gene therapy uses modified viruses (viral vectors) to deliver therapeutic genes to cells.
facultative genes
Genes that are only transcribed when needed
Repressors
Get in the way of the basal transcription factors or RNA polymerase, making it so they can't bind to the promotor or begin transcription
Enhancers
Help the general transcription factors and/or RNA polymerase bind to the promoter
How did Franklins work help Watson and Crick finish their model?
Identified that the x-ray diffraction pattern of DNA was symmetrical and recognized the slope pattern
Sickle Cell Anemia
In patients, the protein subunits that make up hemoglobin stick together to form long, inflexible rods, giving blood cells a sickle-like shape. Sickle cell disease is an autosomal recessive condition that can be caused by mutations in the gene for β-globin (HBB). HBB is one of the two subunits of adult hemoglobin, the protein that carries oxygen in red blood cells. People who inherit two copies of the mutation produce only abnormal HBB that result in sickle-shaped blood cells. The cells get stuck in small blood vessels, leaving body tissues starved for oxygen and causing pain, and, over time, organ damage. Scientists are developing treatments that target the gene switch for fetal hemoglobin. Fetal hemoglobin production is normally turned off soon after birth; keeping it turned on would provide a source of functional hemoglobin in patients.
Meiosis Steps: Detail
Inter I- - DNA replicates Pro I- -CROSSING OVER -PAIRED chromo condense Meta I- -4 chromatids line up MIDDLE cell (metaphase plate) Ana & Telo I- -HOMOLOGOUS separate into HAPLOID Pro II - -sister chromo STAY JOINED to spindle Meta II- Chromo line up single file Ana II & Tela II- -SISTER CHROMO SEPARATE INTO DIFF. HAPLOID CELLS "little squiggles"
How do you get differentiation?
It results from selective gene expression, the turning on and off of specific genes
Leber Congenital Amaurosis
Leber congenital amaurosis (LCA) is a rare genetic disease, affecting 2-3 per 100,000 newborns in the US. It involves the cells of the retina and causes extreme far-sightedness or blindness at birth.
How are genes turned on and off?
Methylation and histone modifications
Polymerase Chain Reaction (PCR)
PCR is a method widely used in molecular biology to make many copies of a specific DNA segment.
histone modification
Proteins bind to histones (proteins that DNA is wrapped around) and causes histones to tighten DNA coiling, which --> turns OFF transcription (Ex. Acetylation)
general transcription factors
Required for the binding of the RNA pol to the core promoter and its progression to the elongation stage, AND Necessary for basal transcription
Rh factor
Rhesus (Rh) factor is an inherited protein found on the surface of red blood cells. If your blood has the protein, you're Rh positive. If your blood lacks the protein, you're Rh negative.
regulatory transcription factors
Serve to regulate the rate of transcription of target genes, AND They influence the ability of RNA pol to begin transcription of a particular gene
What does a repressor bind to?
Silencer
RNAi (RNA interference)
Small interfering RNA (siRN) is incorporated into a protein called RISC. Once the RISC finds a complementary sequence in a mRNA, it cleaves the mRNA in order to destroy or reduce the expression of certain genes. (mRNA Transport)
Small drug molecules
Small-molecule therapies comprise an diverse group of chemical compounds of low molecular weight that are synthesized. Because of their small sizes, these molecules can easily be taken up by cells and may be administered to patients as pills or by injections. (Protein Processing)
Process of Transcription
Step 1: A genes DNA is transcribed by the enzyme RNA polymerase into RNA transcript w/ a complementary sequence
RNA splicing
Step 2: Edits RNA transcript through splicing - a spliceosome (protein complex) trims out introns and splices together exons to produce a mature messenger RNA (mRNA)
mRNA transport
Step 3: Transport from nucleus to cytoplasm
Translation
Step 4: The ribosome translates the mRNA into amino acids, which is brought to the ribosome by a transfer RNA (tRNA), and added to the growing polypeptide chain.
Protein Processing
Step 5: A polypeptide is folded into a 3-dimensional structure and processed to create a mature, functional protein. Processing may involve cleaving, or adding molecules, or assembling two or more polypeptides together.
Primary protein structure
Straight chain of amino acids (Straight/orderly)
How did Chargaffs work help Watson and Crick finish their model?
The 4 bases had complementary ratios
Explain how the change in hemoglobin function affects the structure and function of red blood cells.
The long rods distort the shape of the red blood cells making them sickle shaped. The cells clump together.
Explain how the change in red blood cell structure and function results in the symptoms of sickle cell anemia.
When the cells clump together, they block the normal flow of blood to the limbs and organs causing symptoms such as pain and swelling, infections, and organ damage.
Ribosome
a complex of ribosomal RNA's (rRNA) and proteins that carries out protein synthesis in all cells
CRISPR/Cas9
a unique technology that enables scientists to edit parts of the genome by removing, adding or altering sections of the DNA sequence; when the Cas9-guide RNA complex cleaves the DNA in a cell, the cell machinery will recognize and repair the double-stranded DNA break. Targeting dCas9 to transcription start sites was sufficient to repress transcription by blocking initiation
types of regulatory proteins
activators and repressors
Methylation
addition of a methyl group to a DNA base, or a protein, turns OFF transcription
Huntington's disease
autosomal dominant disease caused by mutations in a gene called huntingtin (HTT), which is required for normal nerve cell function. Mutations in the HTT gene can cause an abnormal protein to be made that causes brain cells to die over time, usually occurs later in adulthood
What is Sickle Cell Anemia?
blood disease that affects the ability of hemoglobin to bind to oxygen.
Evolution
change in allele frequencies in a population many generations
Green fluorescent protein (GFP)
encoded by a single gene; the GFP gene can be used as a visual tag for the expression of other genes.
What did the the double helix structure of DNA confirm?
genetic material must be able to store information; be faithfully replicated and be passed on from generation to generation; and allow for changes, and thus evolution, to occur
reverse genetics
method that is used to help understand the function of a gene by analyzing the phenotypic effects of specific engineered gene sequences. (Genotype known)
Tertiary protein structure
occurs when certain attractions are present between alpha helices and bete pleated sheets
secondary protein structure
occurs when the sequence of amino acids are linked by hydrogen bonds (alpha-helix and beta-pleated sheet)
Quaternary protein structure
protein consisting of more than one amino acid chain
What does positive control refer to?
regulation by activator proteins
transcription factors
regulatory proteins that influence the ability of RNA polymerase to transcribe a given gene
Hardy-Weinberg equilibrium
states that allele and genotype frequencies in a population will remain constant from generation to generation in the absence of other evolutionary influences
What is gene expression?
the appearance in a phenotype of a characteristic or effect attributed to a particular gene.
Gene regulation (general term)
the level of gene expression can vary under different conditions
How was the structure of DNA determined?
the structure of DNA was determined by combining mathematical interpretations of x-ray crystallography data and chemical data.
What does negative control refer to?
transcriptional regulation by repressor proteins
Constitutive genes
unregulated genes, which have constant levels of expression
Chi-square analysis
used to determine whether there is a significant difference between the expected frequencies and the observed frequencies in one or more categories. ... The purpose of the test is to evaluate how likely the observations that are made would be, assuming the null hypothesis is true.
