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12RNA interferenceRNA interference post-transcriptional gene suppression (PTGS)RNAi movie www.nature.com/focus/rnai/animations/index.html3Lecture 19: Drosophila melanogasterPolytene chromosomeLife cycleGenetics/Balancer chromosomeP elements and transformationEnhancer trapGenetic mosaicRead 813-821Read 140-143Fig. D1, D3-D10Fig. 5.25, 5.264Fig. D.15Drosophila melanogaster (fruit fly)Small genome 170 Mb (5% of human genome)33% repetitive DNA13,600 genesThree autosome + X and Y (small chr. number)Giant polytene chromosomeFast life cycle (10 days)6Fig. D.37• Giant polytenechromosomes of larvalsalivary gland are keytools– Replicate 10-11 times– 1024-2048 sisterchromatids stay associatedunder perfect lateralregister– Homologous chromosomestay tightly synapsed– Chromocenter – commonregion where centromerescoalesceFig. 12.158Fig. D.49Fig. D.510Techniques of genetic analysis• In Drosophila, crossing over occurs only infemales– Absence of crossing over in males has considerabletechnical significance• Maintain linkage relationships by inheritance through maleparent– In females, only a moderate amount of crossingover occurs• 1-2 crossovers per meiosis• No crossing over in heterochromatin or 4th chromosome11Balancer chromosomes help preserve linkage• Balancers carrymultiple, overlappinginversions• Most contain adominant marker andrecessive lethalmutation thatprevents survival ofhomozygotes• Useful in geneticmanipulations andmutant screensFig. D.612P-element transposons are critical tools inmolecular genetics• Hybrid dysgenesis– Males from Drosophila strains carrying P elements crossed tofemales that lack P elements– P element becomes highly mobile in germ line of F1 hybrids– Chromosome breakage reduces fertility in hybrids– Progeny of F1 flies carry many new mutations induced by P elementinsertions– Molecular details• P element primary transcript encodes transposase that catalyzestransposition• Cross between P and M strain causes hybrid dysgenesis• Cross between P and P strain does not– Eggs produced by P female have repressor protein that preventstransposition– Repressor coded for by alternatively spliced P element mRNA13Fig. D.71415Transformation: the introduction of cloned DNA into flies• P-elements used asvectors• Insert fly DNA intointact P element andthen into plasmid• Inject into syncytialembryos from M strainmothers• Cross to P males• Mimicking hybriddysgenesisFig. D.8a16Figure D.9P-element as a tool for mutagenizing and tagging genes17Enhancer trapping to identify genes by expression pattern• P element with lacZgene downstream ofpromoter• When mobilized, 65%of new insertionsexpress lacZ reporterduring development• Promoter can onlyactivate transcriptionif under control ofenhancers of genesnear insertion site• Detects genes turnedon in certain tissues• Genes isolated byplasmid rescueFig.


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UMD BSCI 410 - Lecture 19: Drosophila melanogaster

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