Genetics 4 Flashcards
Analysis of nucleic acids - goal is what
To find defective genes or abnormal gene expression
Analysis of genes includes what
Locating a disease gene - require intense research
Diagnosing the presence of a disease gene - routine task
Analysis of gene expression is what
Quantifying RNA
Polymerase chain reaction (PCR) is used for
Amplification of DNA fragments
PCR can be used how
With allele specific oligonucleotides that bind only to mutant or wildtype alleles OR
Can be used to know if there are insertions or deletions by counting the length of the amplified fragment
Reverse Transcriptase PCR
PCR only works with DNA so if you want to analyze RNA you have to transcribe the RNA into DNA
Reverse Transcriptase PCR - In a first step…
Reverse transcriptase is used to transcribe RNA into cDNA
Reverse Transcriptase PCR - second step
cDNA serves as a template for a PCR reaction
Reverse Transcriptase PCR - the more RNA there is,
the more DNA will be amplified
Quantitative PCR - is what
Accurate quantification of specific DNA molecules
Amplification is monitored in real time
Quantitative PCR - Often used for
mRNA detection
copy of mRNA into cDNA first, then start PCR
Allele specific oligonucleotides are designed for what
To bind just one specific allele
Allele specific oligonucleotides - hybridization of genomic DNA to ASO demonstrates
presence of the allele
Allele specific oligonucleotides - what are the three possible outcomes
1 All of it binds to wildtype spot and non to mutant (homozygous for wildtype)
2 All of it binds to mutant and none to wildtype (homozygous for mutant)
3 Half to wildtype and half to mutant (your patient is heterozygous)
Sequencing
Reads it base by base and forms the nucleotide sequencing
Can be used to determine if a patient is heterozygous at a specific locust
What to sequence
Whole exome sequencing
SNP typing
Only sequence SNPs that matter
RNA
What to sequence - Whole exome sequencing
All of the genome that is expressed
30 million base pairs
What to sequence - SNP typing
Only the bits of the genome that vary
Less than 10 million base pairs
What to sequence - Only sequence SNPs that matter
A few thousand
What to sequence - RNA shows
Variants and gene expression profile
Prenatal Diagnosis - Sampling techniques
Amniocentesis
Chorionic vilus sampling
Prenatal Diagnosis - Sampling techniques - Amniocentesis
Gold standard
Amniotic fluid is withdrawn with a needle at 15/16 weeks gestation
Fluid contains fetal skin cells
Prenatal Diagnosis - Sampling techniques - Chorionic Vilus Sampling
Cells taken from chorion with a needle at 10-12 weeks gestation
Chorionic cells are extraembryonic in origin
Chance of missing a mosaicism with this
Preimplantation diagnosis
Can be performed on early stage embryos
One cell can be removed from embryo and analyzed
Used to select embryos for implantation
Suitable for diagnosing Mendelian and chromosomal disorders
Legal restrictions to address ethical concerns
Direct to Consumer Genetic Testing
23 and me
Can buy the kit and they look at the few thousand SNPs that are related to disease
Before you test - think about
What will you do with the info
Will it help prevent disease or just worry the patient
Who will have access to the info
What do you do about incidental findings
HIPPA
Protects the privacy of medical records, including genetic data that may be part of such record
GINA
Genetic Information Nondiscrimination Act - no individual can be discriminated against based on genetic information
This is getting fought right now though
