Translational Research, DNA Analysis/Tech Flashcards
Understand principles behind various molecular techniques
What are restriction endonucleases?
DNA endonucleases that recognize spec palindromic DNA seq’s
What do restriction enzymes produce?
- sticky ends that can be ligated together
Why is DNA hybridization used?
To ID presence of a target seq using a complementary DNA probe
Describe in situ hybridization
- probe used to assess expression of gene in a spec tissue, or at diff embryologic stages
What is cDNA used for?
Creating templates for microarray analysis
Purpose of microarray analysis?
- allows simultaneous measurement of every gene in the pop for its expression pattern
Main objective of PCR?
- amplify a target fragment using antiparallel complementary seq-spec primers that flank amplified target
What do Restriction Fragment Length Polymorphisms (RFLPs) do?
- polymorphisms in genome that either create or destroy a restriction enzyme site
Practical use of microarrays?
- allows for real-time analysis of effectiveness of drug tx of spec cancers
- measures not just presence of gene but also level of gene expression
3 major characteristics of DNA polymorphisms
- genomic variations in a gene w/i the pop (at least 1%)
- may involve coding or noncoding regions
- maintained (eg sickle cell)
How to detect RFLPs?
Southern analysis
What are allelic-spec oligonucleotide probes?
PCR or hybridization using allele-spec oligo
How to use repetitive DNA to detect polymorphisms?
- Variable Number of Tandem Repeats (VNTR)
- digestion flanking VNTR region
- DNA fingerprint every ind
- detect family relationships or condemn/acquit suspects
How does sickle cell disease affect spec sites in gene seq?
- normal gene has MstII restriction site
- sickle cell gene has no MstII site
- this can be taken adv of in RFLP & Southern blot analysis
What is a therapeutic application of recombinant DNA tech?
- vaccines for infectious antigens (minus the agent itself)
