Genomics Exam 1
What is cloning?
An exact duplication of DNA
List the ways nucleic acids can be chemically mutated
-Methylation of bases -Deamination of bases (5-methylcytosine to thymine and usually goes uncorrected)
Define the term polycistronic
-Multiple genes, especially if needed expressed at the same time -To do this a SNP would be located in the promoter region
How many chromosomes are in humans?
-22 pairs of autosomes; one pair of sex chromosomes (X, Y) -23 pairs or 46 chromosomes in "normal" somatic human cells
What is p53 and why are mutations to this gene and protein correlated with a higher cancer rate?
-A DNA binding protein that recognizes DNA damage and signals to stop the cell cycle from entering S phase -P53 mutations permit cancer to grow and have resistance to chemotherapy and radiation
What is a gene?
-A DNA segment that contributes to a phenotype/function -Codes for an RNA product
Genomic Markers
-A measurable DNA and/or RNA characteristic that is an indicator of normal biologic processes, pathogenic processes, and/or response to therapeutic or other interventions -Can measure expression, function, regulation
What is PCR?
-A method to duplicate DNA in the test tube -Real time PCR: a method that used a fluorescent dye to indicate when enough DNA double helical copies have been made
What are multigene proteins?
-A protein that has multiple subunits from multiple genes -Hemoglobin has an alpha and a beta gene, plus a gamma gene expressed during development and a delta gene that upregulates at high altitude
Lynch Syndrome
-Also called hereditary non-polyposis colorectal cancer (HNPCC) -MLHL, MSH2, MSH6, PMS2, EPCAM
Explain how mutations to the Shine-Dalgarno sequence could affect expression?
-Can decrease protein synthesis for a gene -This would prevent the RNA from being translated into a protein
What is Favism?
-Caused by a glucose-6-phosphate dehydrogenase deficiency, which is a genetically inherited trait -Fava beans contain divicine which produces H2O2 which can cause hemolytic anemia in G6PD deficient individuals
How are genetic tests done?
-Clinical setting and home kits -Sample types of blood, saliva, cheek cells
Non-Small Cell Lung Cancer (NSCLC)
-Crizotinib: kinase inhibitor -Indicated when anaplastic lymphoma kinase (ALK) or ROS1-positive
What information is available with genetic testing?
-DNA sequence, amino acid sequence, SNPs, possible disease likelihoods, possible unique reactions to drugs -Can test stuff like lactose intolerance, caffeine consumption, sickle cell anemia, tay-sachs, Zellweger syndrome -Can search for variants in a patient's DNA
How do you get G6PDD?
-Defect of the enzyme glucose-6-phosphate dehydrogenase -X-linked recessive (mostly males get it from mother who was either carrier or had disorder, females only get it if their father had the disorder and their mother was at least a carrier) -Caused by a SNP in an exon
Clinical Pharmacogenomics
-Developing processes for: ordering, reporting, and interpreting pharmacogenomic tests -Testing can be preemptive (proactive) or reactive
What is the effect of consanguinity on recessive disorders?
-Doubled if parents are first cousins to about 3-5% -High increase with inbreeding
Biomarkers examples
-Examples: DNA sequence, gene expression -Specific biomarker: prostate specific antigen (PSA)
What is G6PDD?
-G6P is part of the oxidative phase of the pentose phosphate pathway -It produces 2 NADPH so if there is a G6PDD then we get no NADPH -NADPH is the essential molecule needed by the red blood cell that is only available to those cells from the pentose phosphate pathway
Autosomal dominant diseases
-Huntington's disease -marfan's disease
Variation is the rule
-Is the RULE not the exception -Essential for survival -Some are disadvantageous, some advantageous, some neutral
Mitochondrial disorders
-Leber's syndrome -leigh syndrome -mitochondrial myopathy
Cystic Fibrosis
-Mutation G551D in CFTR gene leads to impaired chloride channel function -Ivacaftor indicated for patients with CFTR G551D mutation
Why do mutations to signaling systems cause both metabolic concerns and vulnerability to cancers?
-PKA mutations: lead to altered expression -RAS: leads to cancer
What are histones? What are the subunits? What role do they play in the chromosome?
-Proteins that bind DNA -8 histone subunits in the histone core (H2A, H2B, H3, H4) and 1 holding the DNA to the histone core (H1) -This makes up the structure and composition of a chromosome
What is genetic anticipation?
-Severity of dominant mutations increases through generations -Ex: Huntington's disease
What is DNA fingerprinting?
-Southern blots are a way to look at only pieces of interest with DNA probe -Unique RFLP pattern for a specific DNA
Describe how to make a cDNA
-Take RNA from the cell and mRNA reflects sequence of gene without introns, only looking at exon portion of the gene (this is the cDNA) -Can define where in the chromosome we have the gene
What is the largest pharmacogenomics consideration a pharmacist must make?
-The difference between males and females -Estrogen not given to men, testosterone not given to women
Why would mutations in the mismatch repair (MMR) enzyme system be associated with higher cancer rates?
-The loss of MMR allows for a significant number of spontaneous mutation to go uncorrected. -A significant number of cancers are associated with mutations of enzymes in these pathways. -Specifically, nonpolyposis colorectal cancer is associated with mutations in these pathways
X-linked dominant diseases
-fragile X syndrome -rett syndrome
X-linked recessive diseases
-hemophilia -fabry disease -G6PPD -lesch-nyhan
Prader-Willi Syndrome
-mutation in proximal part of chromosome 15 coming from the father -due to no expression of gene 2
Angleman Syndrome
-mutation in proximal part of chromosome 15 coming from the mother -due to no expression of gene 1
Autosomal recessive diseases
-sickle cell -cystic fibrosis -tay-sachs
Y-linked dominant diseases
-wyer syndrome -Y chromosome infertility
In humans, how many chromosomes do we have?
23 pairs, 46 total, during S phase we have 92 chromosomes going into M phase and then separate into the daughter cells
Histone Core
8 histone proteins
What are microarrays
A chip that has DNA sequences covalently attached and can be used to evaluate which genes are expressed
How do the bases pair up in the double helix and how does the cell know when a mismatch takes place?
A:T (11 A) and C:G (10.8), other combos can hydrogen bond but the distance will be different and be recognized as incorrect
Pharmacogenomics
Defined as a field of research that studies how a person's genes affect how that person responds to medications
A nonsense mutation
AKA early termination mutation; caused by the creation of stop codon before completely reading the code for a protein
Hereditary breast cancer
BRCA1 and BRCA2
How is the DNA analyzed?
Break the cells open, cut up the DNA with specific endonucleases, type II restriction endonucleases are used for these techniques
What is the molecular basis of this disorder?
Change in AA creates problems and creates defects in enzymes
What are trisomys?
Change in chromosome number/size causes disability/disease
What is mtDNA?
Circular, commonly have more than one nearly identical DNA molecule, can divide inside of the cell, can merge together to make larger mitochondria
Ortholog (Orthologue)
separated by a speciation event; same protein in different species
How are chromosomes condensed, especially when that are inactive?
Coil up histones and make them into loops, fold the loops and creates thickness of chromosome, especially during meta phase prior to cell division when DNA is doubled in number
How mutations in histone, methylases, and acylases affect gene expression?
DNA is negatively charged, histones are positively charged. Methylation and acylations of histones neutralizes the charges on histones and regulates the release of DNA from those histones
Why does Favism cause hemolytic anemia?
Fava beans can interfere with the glutathione reaction which inhibits the ability to reduce and eliminate peroxides which reacts with the double bonds in the unsaturated fatty acid tails of the membranes, making the membranes rigid and easy to break causing hemolysis
Paralog (Paralogue) Examples
Heme containing proteins of cytochrome P450 and myoglobin
Ortholog (Orthologue) Examples
Human insulin and bovine insulin
Homolog (homologue)
Sequences or structures that are related to each other by descent from a common ancestral sequence or structure are homologues. All members of a gene family are homologues
Li-Fraumeni Syndrome (LFS)
Mutations in TP53 gene, CHEK2 gene
Why would mutations to introns affect protein synthesis?
Mutations in these sequences may lead to retention of large segments of intronic DNA by the mRNA, or to entire exons being spliced out of the mRNA. These changes could result in production of a nonfunctional protein.
How does X-inactivation affect symptom level in females?
Occurs very early in life, inactivation is random, wide implications for X-linked disorders
How do we know that DNA is the molecule that carries genetic information?
Phosphate containing molecule and not a sulfur containing molecule. Phosphorus is a permanent component of DNA, but only a transient component of Proteins. Sulfur is a permanent component of proteins but sulfur is not part of the structure of DNA
Which drugs should not be given to a patient that is G6PDD?
Primaquine
What is the "upstream region" of a gene?
Promoter/enhancer regions that can change expression and change concentration of protein product with no effect on the activity of the protein
Mutations in JAK2 (V617F)
Risk of essential thrombocytopenia
What are introns?
Sequencing of DNA that are not a part of our genetic code they are inserted between segments of exons, made up of DNA similar to viruses and must be removed to express proteins, some self-splice some need removed by snRNPs
What is alternate splicing?
Splicing genes to give more than one protein
How is genetic testing done?
Take a sample from blood or salvia
cDNA
complimentary DNA, made by reverse transcription of mRNA (mature) to create a cDNA library
GAA -- GAC
conservative missense mutation
Phylogeny
The evolutionary history of a molecule (such as a gene or a protein), or even a species and the relationship between such entities
TGA -- TGG
run on non-sense mutation
Explain how mutations to the upstream regions of a gene, or the associated proteins affect gene expression
The upstream regions contain the promoter and enhancer sites
How does mutations in topoisomerases, methylases, and acylases affect gene expression?
Topoisomerases are needed to unwind DNA
Biomarkers
Used as an indicator of a biological state
What are restriction endonucleases?
Used to chop up (restrict) foreign DNA
Point mutation
a change of one location
Conservative mutation
a change to the gene product causes only a small effect such as switching a glutamate for an aspartate, both are acid amino acids
Duplication mutation
a piece of DNA is abnormally copied one or more times
SNP
a single change to a nucleic acid base
Mutation
a variant that occurs less than 1% of the time in the population
Nucleosome
about 200 base pairs of DNA and 9 histone proteins
Insertion mutation
addition of one or more nucleic acid bases to the DNA sequence
Histones
basic proteins that bind DNA
SNP in a stop codon
cause a run on non-sense mutation
A run-on mutation
caused by a mutation in a stop codon leading to a longer than normal protein
SNP in an exon could
change an amino acid or introduce an early stop codon
A missense mutation
changes an amino acid
SNPs in the upstream region could
decrease expression of the protein
GAA -- GA
deletion, frame shift non-sense mutation
CAG -- TAG
early termination nonsense mutation
Trisomy 21
extra copy chromosome 21- Down's syndrome (usually from mother)
Trisomy X
extra copy of X chromosome in females: behavioral defects, learning disabilities, delayed motor skills
XXY
extra copy of X in males: sterility, gynecomastia
XYY
extra copy of Y in males: hyperactivity, learning difficulties
Trisomy 13
extra copy of chromosome 13- severe retardation, death, hand deformity
Trisomy 18
extra copy of chromosome 18 - microcephaly, clenched hands and overlapping fingers
SNP at an exon-intron junction could
fail to remove an intron and cause a non-sense mutation
CNV
gene that occur more than once on the same chromosome in the genome and sometimes change in the number of copies
"Reverse genetics"
identification of atypical amino acids helped to identify mutations at the DNA and mRNA level which lead to different drug responses
GAA -- GAAA
insertion, frame shift non-sense mutation
Deletion mutation
loss of one more nucleic acid bases to the DNA sequence
Frameshift mutation
loss of reading frame due to an insertion of deletion mutation
Paralog (Paralogue)
separated by a gene duplication event; similar proteins in same species
Wild-type
most common allele in a population
De Novo Mutation
new mutations that occurred in this individual
DNA
no oxygen at position 2 of ribose ring, uracil is replaced by thymidine
Silent mutation
no physical change is created
GAG -- GTG
non-conservative missense mutation
Inherited Mutation
occurred in a previous generation and was passed to the offspring
Centromere
point of attachment of duplicated chromosome after S phase until separation in M phase
SNP at an intron-exon junction could
remove an exon between two introns
Repeat expansion
repeated sections of DNA increase the number of repeats
Telomeres
repetitive DNA caps on the ends of chromosomes to protect them from degradation
VNTR
repetitive sequences of DNA outside the gene
Microsatellites
short repeats of less than 5 bases that occur in the DNA
CTG -- TTA
silent mutation
What is genetic imprinting?
the genetic defect differs when it comes from one parent verse another
Heterozygous:
the pair of chromosomes has different alleles
Homozygous
the pair of chromosomes has the same alleles
Allele
the specific version of a variable form of the same gene
RNA
there is an oxygen at position 2, and no methyl group at position 5 when using uridine
Polymorphism
two or more variations of the same DNA section
Sequence homology
two sequences that came from the same ancestral sequence
Genomic Library
using radio labeled cDNA probes to identify which fragments contain a gene
