Post on 30-Sep-2020
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How can PCR be used to mutagenizeDNA or to introduce novel sequence into a DNA fragment ?
Presented by: Marguerite Nelson,Xuejing Zhang ,Yanna Ding
How can PCR be used to mutagenize DNA or to introduce novel sequence into a DNA fragment?
Allows scientists to study specific mutations in DNA and their phenotypic expressionCan create mutant animals to study mutation effect in animals instead of humans
How can PCR be used to mutagenize DNA or to introduce novel sequence into a DNA fragment?
Mutate gene by site-specific in vitro mutagenesis----using PCRthe point : using siutable primers containingmutagenic information
Figure from Essential iGenetics
Two reactions
Primer 1 and 1M product is A
Primer 2 and 2M product is B
Combine A and B
Denature
Reanneal
Elongation
Run PCR with primers 1 and 2
Figure from Principles of Biochemistry
Then,check the expressionof mutation
What are other uses for PCR besides merely amplifying DNA?
Application in gene-related research
Amplification of specific single RNA molecule from mixtureBuilding cDNA libraries by RT-PCR Quantitation of DNA or RNA by Real-time quantitative PCRDNA sequencing Detecting mutations Making labeled probes to screen libraries or carry on blotting experiments
Human Health• Detecting bacterial and viral infections. • Disease diagnosis.• screening for human genetic disease base on
the analysis of microsatellites as a screening marker.
• Eventually play a role in identifying individuals with cancer susceptible genes
• Monitoring cancer therapy • Sex determination of prenatal cells or prenatal
diagnosis.
The Law
• Forensic fingerprinting from collected evidence
Modern Systematics, Ecological Studies, and Animal Behavior
• Glean genetic information from the faintest traces of the shyest, rarest animal-urine, feces, scent marks, infinitesimal bits of hair or skin rubbed onto a tree as the elusive creature passes by.
Evolutionary Relationships
• Sort out relationships among vanished human groups, and tracing human migrations
Reference• P.J.Russell, Essential iGenetics(Benjamin Cummings, San
Francisco,2003),pp.405-407.• D.L.Nelson,M.M.Cox, Lehninger Principles of
Biochemistry(W.H.Freeman and Company, New York,ed,H,2005)pp.316-317.
• B.D Hames & N.M Hooper, Instant Notes in Biochemistry (Second Edition)(BIOS Scientific Publishers Limited,2000) pp.263-265.
• P.C Turner,A.G.McLennan, A.D. Bates&M.R.H.White, Instant Notes in Molecular Biology(Second Edition)(BIOSScientific Publishers Limited,2000) pp.89-92,174-177.
• Bruce Korf, New England Journal of Medicine(MolecularDiagnosis,1995) pp.1499-1520.
• Norman Arnheim, Tom White & William E. Rainey, BioScience(Application of PCR: Organismal and Population Biology, 1990) pp. 174-182.