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Lab Practical DNA Fingerprinting notes kcfac kilgore cc tx us gel
Biology 1408 Laboratory Inquiry–Based DNA Technology Laboratory DNA Fingerprinting Notes I. Learning Outcomes A. Student Should Be able to: --identify virus and viral infection --Identify hemorrhagic fever --Identify agarose gel --Identify electrophoresis --Identify restriction digest analysis --Identify DNA fingerprinting analysis B. The student must demonstrate successful completion of the laboratory exercise: Read Student Worksheet --cast agarose gel (20 pts) --load gel (20 points) --electrophoresis (20 pts) C. The student must submit results and conclusion --Describe the results: Compare the fingerprints of the Pennsylvania, Alabama, and Missouri virus strains. (20 pts) --What are your conclusions about the virus infecting the 3 Missouri patients (20 pts) II. What is the nature of viruses and viral infection? --viruses were first discovered by microbiologists seeking the cause of a plant Disease --a nonliving, complex molecule --infectious particles, consist a protein coat ,containing nucleic acid of RNA or DNA --protein coat on is called a capsid, encloses the genome --some viruses also have an outer envelope --Viral Infection --viruses require a host cell to reproduce --inserts viral genome into that of host, transforming the normal cell into a cancer cell --virus may remain lysogenic (dormant) or lytic, death of host cell 1 --Virus and cancer --some groups can cause cancer: Retroviruses, papovavirus, adenovirus, ands herpesvirus --Emerging viruses --are not new but evolving, expanding into new species territories --sudden appearance in the late 1970s. --1976, Ebola in Ziare (central Africa) --virus caused hemorrhagic feve --often fatal, fever, vomiting, bleeding, collapse of circulatory System. 4 decades this virus went unnamed. --1993, dozens died from hantavirus infection, southwestern U.S. --1995, Ebola (acute hemorrahagic fever) --central Africa, Zaire --1999, encephalitis, the Nipah virus in Malaysia, killed 105 people, destroyed the pig industry --2003, SARS (severe acute respiratory syndrome --pneumonia-like symptoms --natural host are a weasel-like mammal, Civets in China --other wild animals --meat is eaten as a delicacy --AIDS (acquired immune deficiency syndrome) --caused by the retrovirus, HIV --Influenza --natural hosts are birds, migratory --new strains emerge each year --millions affected --possibly deaths III. Three factors contribute to viral emergence --mutation of an existing viral strain --mutation of a strain which can cross between host species --exposure of an unknown viral agent to a larger area IV. Basic health measures to prevent viral outbreaks --water purification --mosquito control --secure sewer systems V. The use of biotechnlogy in the separation of macromolecules for identification A. gel electrophoresis is used --to separate molecules on basis of size and charge (+/-) --in forensic laboratories to help in solving crimes --to identify contagious and possibly deadly agents 2 VI. How gel electrophoresis works A. charged macromolecules migrate through a gel when placed in an electrical field --smaller molecules migrate faster than larger ones VII. What is the nature of the gels used in electrophoresis A. the gels contain the complex carbohydrate, agarose --extracted from seaweeds (red and brown algae) --a linear polymer B. the powered agarose is mixed in a buffer containing water, melted by heating To boil until clear (10-20 minutes). --allow to cool to 65oC before use C. the gel forms a matrix (contains small pores acts like a sieve). IX. Restriction enzymes A. produced by bacteria --cut DNA molecules into smaller fragments --each enzyme recognize a specific restriction site --most restriction sites are composed of 5-6 base pairs B. the restriction Eco RI was isolated from E.coli --recognition sequence : ...………GAATTC……….. …………CTTAAG………. ............G^AATTC……. ………CTTAA^G……. X. DNA Fingerprinting A. DNA molecules from all living organisms and viruses contain restrictions sites B. Restriction analysis can be used to identify a specific organism. C. Restriction analysis of specific sequences of DNA from different organisms produce: --different size restriction fragments D. Small DNA fragments and viral genomes can be cut with restriction enzymes --electrophoresis produce a recognizable pattern --the patterns produced can be used to identify differences in DNA sequences of organisms 3
"Lab Practical DNA Fingerprinting notes kcfac kilgore cc tx us gel"