| SIU School of Medicine - How can PCR be used to mutagenize …bbartholomew/PCR.pdf · 2007. 9....
Transcript of | SIU School of Medicine - How can PCR be used to mutagenize …bbartholomew/PCR.pdf · 2007. 9....
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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
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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
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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
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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
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Figure from Principles of Biochemistry
Then,check the expressionof mutation
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What are other uses for PCR besides merely amplifying DNA?
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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
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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.
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The Law
• Forensic fingerprinting from collected evidence
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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.
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Evolutionary Relationships
• Sort out relationships among vanished human groups, and tracing human migrations
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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.
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