BOLOGY 107 Lecture 37 Outline of Last Lecture I Viruses a HIV II Viroids and Prions a Viroids b Prions Outline of Current Lecture III Biotechnology a DNA cloning b Other DNA techniques Current Lecture Biotechnology 1 DNA cloning a Making multiple copies of DNA b Recombinant DNA i Combines DNA from many sources c Plasmids i Circular extra chromosomal DNA ii Can b made to carry foreign DNA iii Used for gene cloning d Restriction enzymes i Isolated from bacteria 1 Meant to protect from foreign DNA and viral infection 2 Cleaves DNA at specific sequences a Restriction sites b Leaves sticky sites with unique complementary sequences 3 Any restriction fragments with compatible ends can be glued together with ligase e Process i Isolate pieces of DNA 1 Cut with restriction enzymes 2 Separate by size on gel cut up and purify ii Ligate to form plasmid These notes represent a detailed interpretation of the professor s lecture GradeBuddy is best used as a supplement to your own notes not as a substitute iii Transform into bacteria iv Grow lots of bacteria make many copies of the plasmid f Why do it i Subsequent cloning and DNA manipulation ii Have bacteria make a specific protein 1 Used for studies or drug production g Libraries i Collection of different plasmids viruses or artificial chromosomes that together represent the whole genome of an individual 1 Can be genomic or cDNA 2 Stored in 96 well plates h cDNA i DNA copy of mRNA 1 RTase used to copy mRNA to DNA 2 DNA polymerase used to make copies 3 Can make a library using either the primer or just one specific mRNA 2 Other DNA techniques a PCR i Rapid and efficient DNA amplification ii Used to copy specific DNA iii Based on DNA replication 1 Uses Taq polymerase from Thermos aquaticus a thermal Archaea iv Three steps repeated 1 Denature the DNA separate with heat 2 Annealing allow synthetic primers to bond while cooling 3 Extension polymerase makes copies 4 Can be started at one sequence or done with the whole genome v Uses 1 Diagnosis detect virus or disease causing allele 2 Forensics amplify DNA from evidence 3 Environmental monitoring identify species in soil or water 4 Gene expression measure changes in mRNA levels 5 Genetic manipulation isolate for cDNA cloning a Evaluate mutation effect b Distinguish alleles vi Problems 1 Taq polymerase has no proofreading ability a High error rate 2 It can also detect all DNA including contamination
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