BSCI410 -What types of changes in the DNA

Name:UID:Homework II, BSCI410 Fall 2016 (100 points)Due October 25 in class (paper preferred)Please write neatly!You have done a genetic screen using the mutagen ethylmethane sulfonate in abeneficial species of insect to identify genes that will allow it to survive betterunder dry conditions. You used visible markers to map the genes on its 5chromosomes. The questions below are independent of each other but all relateto this genetic screen. Assume that this species is a standard diploid, that youcan identify males and females cross them to each other, and that you havemethods to make transgenic animals. (Note that this species and screen arepurely hypothetical so you won’t be able to find the screen that Dr. Pick has inmind by a google search)1. (5 points) What types of changes in the DNA were generated in your screen?Justify your answer in one sentence. 2. (15 points) Among the mutants you identified, 6 of them that map to the samechromosome show remarkably similar phenotypes. There are very few visiblemarkers on this chromosome so you cannot tell how many genes arerepresented by these 6 mutants, named a,b,c,d,e,f. You decide to carry out acomplementation test. You figure out that mutants b and e map to the samegene, while all the others map to unique genes. Draw a table that woulddemonstrate these results (hint: the Hartwell book shows a complementationtest). 1 Name:UID: 3. (15 points) You have obtained sequence of three of the genes and theyappear to encode transcription factors. You have cloned each of them and candetect their expression by in situ hybridization. Gene S is expressed in a largeregion in a circular shaped “water retention” tissue, while gene T and gene V areexpressed in the anterior and posterior half of this tissue, respectively. Youdetermine that Gene S is required to activate genes T and V and that gene Tcompletely represses gene V expression. Draw the expression of these genes inanimals: (Note: it may be helpful to use different colors for each gene in yourdrawings, please define which color represents which gene).A) Carrying a deletion of gene S. B) Carrying a deletion of gene T C) Ectopically expressing gene T under the control of a gene S enhancer 4. (15 points) You would like to determine the cis-regulatory sequences thatdirect gene S expression in the “water retention” tissue. You make use of theDrosophila P-element containing plasmids and a GFP gene to do this andidentify an enhancer in the intron of gene S.A) (8 points) Using diagrams similar to those shown in the answer toChapter 17 Problem 19, draw the constructs you would have made to showthis, labeling each of the parts that are labeled in that figure. 2 Name:UID: B) (4 points) In two sentences only, how would you make use of havingthis reporter gene to genetically identify regulators of gene S? C) (3 points) Name one assay you could do to determine whethercandidate regulators of gene S are direct transcriptional regulators. 5. (15 points) The following questions apply to one mutant, “r” that appears toencode a transcription factor. Gene r mutants can live for 3 days with nowater (normally, this species requires water every day). You hypothesize thatthis mutation is a gain-of-function mutation.A) Using simple line drawings, diagram the DNA fragments you wouldhave to insert into transformation vectors to generate insects to test thishypothesis using the UAS-GAL4 system. Do not draw all the components ofthe transformation vector. B) Diagram the crosses you would do. Note: you do not need to useBalancer chromosomes for your crosses. C) State in one sentence the expected outcome that supports yourhypothesis. 3 Name:UID:6. (15 points) Although in real-life, insects have not been shown to imprint theway mammals do, for this hypothetical case, one of the genes identified in yourscreen, gene Q, appears to be paternally imprinted. Draw the crosses (withgenotypes and phenotypes) that would have led to this conclusion. 7. (20 points total; each answer is worth 1 point unless otherwiseindicated)You clone and sequence one of your genes and do a blast search. You find agood match to Drosophila oatp58Db. To learn more about the potential functionof this gene, you log on to Fly base (http://flybase.org/).Please actually go to Fly Base and enter oatp58Db into the Quick search, thenclick on genes, and then Oatp58Db to answer the following questions:A) What is the full name of this gene?B) Based on Gene Ontology (GO) what is the molecular function of theprotein encoded by this gene?C) Based on the information given here why would you want to do furtherexperiments before being sure of this function?D) Move down the page to “Summaries.” There are 5 alleles reported.How many of these alleles could have been isolated in a mutant screen, such asthe one described in this Homework?E) Move down to Expression Data, High-Throughput Expression Data,FlyAtlas Anatomy Microarray. What tissue and stage is this gene expressed in?F) Open modENCODE Anatomy RNA-Seq. What tissue is expressionseen using this technique?G) How might you explain the discrepancy between E and F? 4 Name:UID:H) Open modENCODE Developmental RNA-seq. What is the earlieststage this gene is expressed?I) Open modENCODE Treatments RNA-seq. What treatment moststrongly induces expression of this gene?J) Moving down the page, name one human ortholog of this gene.K) One human ortholog has an “OMIM” entry. Follow the link to this entry.What is the syndrome? L) This entry suggests redundancy in humans. Explain. (3 points) .M) How could you use Drosophila to test whether the two human genes,SLCOB1 and SLCOB3, each have the same basic function as the fly gene? (3points) N) Return to Flybase. Open Gene Model and Products. How many exonsare indicated for oatp58Db?O) Comparing this to the CDS (Coding DNA Sequence), what can youconclude? (2 points) 5

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