You have identified an enhancer that drives Pax6 expression in the developing lens and cornea of the mouse eye. To identify regions of the enhancer that bind lens specific transcription factors (transcription factors expressed only in the lens but not the cornea) you perform a linker scanning experiment in which you mutate 10 bp regions centered around positions 5 to 95 bp of the enhancer. The enhancer variants are tested for their ability to activate expression of Pax6 in both the cornea and lens of the developing mouse. You isolate each tissue separately and carry out a northern blot for Pax6 mRNA on each sample as shown below. List which lane A-J of the northern blot contains a region of the enhancer that binds a lens-specific transcription factor. Explain how you determined this referencing specific data from the gel and your knowledge of the molecular biology involved in regulating transcription. A B C D E F G H I J 5 15 25 35 45 55 65 75 85 95 WT Cornea Lens Northern Blot for Pax6 mRNA

You have identified an enhancer that drives Pax6 expression in the developing lens and cornea of the mouse eye. To identify regions of the enhancer that bind lens specific transcription factors (transcription factors expressed only in the lens but not the cornea) you perform a linker scanning experiment in which you mutate 10 bp regions centered around positions 5 to 95 bp of the enhancer. The enhancer variants are tested for their ability to activate expression of Pax6 in both the cornea and lens of the developing mouse. You isolate each tissue separately and carry out a northern blot for Pax6 mRNA on each sample as shown below. List which lane A-J of the northern blot contains a region of the enhancer that binds a lens-specific transcription factor. Explain how you determined this referencing specific data from the gel and your knowledge of the molecular biology involved in regulating transcription. A B C D E F G H I J 5 15 25 35 45 55 65 75 85 95 WT Cornea Lens Northern Blot for Pax6 mRNA

Human Anatomy & Physiology (11th Edition)

11th Edition

ISBN:9780134580999

Author:Elaine N. Marieb, Katja N. Hoehn

Publisher:Elaine N. Marieb, Katja N. Hoehn

Chapter1: The Human Body: An Orientation

Section: Chapter Questions

Problem 1RQ: The correct sequence of levels forming the structural hierarchy is A. (a) organ, organ system,...

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The UG4 gene is expressed in the stem and leaf tissue of the plant Arabidopsis thaliana. To identify potential mechanisms regulating UG4 gene expression, six small deletion mutations are made in cloned sequence containing the upstream regulatory region. The full length mutant segments Which mutation(s) affect an enhancer? Why? Which mutations identify the promoter? Why? Speculate about the reason for different transcription rated obtained for fusion constructs E and F.
a. Would you expect a cell to continue or to stopdividing at a nonpermissive high temperature if itis a temperature-sensitive Ras mutant whose protein product is fixed in the GTP-bound form atnonpermissive temperature?b. What would you expect if you had a temperaturesensitive mutant in which the Ras protein staysin the GDP-bound form at high temperature?
A cloned gene fragment contains a regulatory element that isrecognized by a regulatory transcription factor. Previousexperiments have shown that the presence of a hormone resultsin transcriptional activation by this transcription factor. To studythis effect, you conduct an electrophoretic mobility shift assayand obtain the following results: Explain the action of the hormone.
Using RNA-seq analysis of transcripts isolated from different organs in mice, you find that the GFAPgene is expressed only in the brain. You examine expression in glia and neurons (two different celltypes found in the brain) and determine that GFAP mRNA and protein is made only in glia. Thisgene has never been studied before, so you start by characterizing the cis-acting elements requiredfor brain-specific expression. a. You decide to fuse pieces of DNA found upstream regions of the coding region of GAP to areporter gene (lacZ) and monitor expression. Different fragments (shown as dark lines) werecloned next to the lac gene that lacked its own promoter. M and H are restriction enzymesites. From the data shown, determine which region contains the promoter and whichregion contains an enhancer . Explain.
The transcription of a gene called YFG (your favoritegene) is activated when three transcription factors (TFA,TFB, TFC) interact to recruit the co-activator CRX. TFA,TFB, TFC, and CRX and their respective binding sitesconstitute an enhanceosome located 10 kb from the transcription start site. Draw a diagram showing how youthink the enhanceosome functions to recruit RNA polymerase to the promoter of YFG.
Enhancer RPA Gene A Use the diagram above, which depicts a chromosomal region less than 500,000 bp, to determine the expression of each gene under the scenarios specified in the table. Regulatory promoters are indicated by "RP". Genes A, B, and C have intermediate expression when no regulatory proteins are bound. Complete the table by choosing high, intermediate, or low to describe the expression level of each gene under each specified scenario (i and ii). No regulatory proteins bound (i) A repressor binds to RPA and an activator binds to RPC (ii) A regulatory protein binds at each of the activator, insulator, and silencer. Expression of A? Intermediate 1. [Select] 4. [Select] Insulator RPB Expression of B? 2. Gene B Intermediate 5. [Select] [Select] Silencer RPC Expression of C? 3. Gene C Intermediate 6. [Select] [Select]
In Figure 13-5, two different methods are illustrated forvisualizing gene expression in developing animals.Which method would allow one to detect where within acell a protein is localized?
In the module, you have learned about P-element mediated transgenesis in Drosophila and the concept of using transgenes to rescue mutant phenotypes. In the figure below, you will see a wild type fly with its natural eye colour and three mutants with their eye colours changed to vermillion, white and rosy, respectively. A schematic of P-element mediated transgenesis (as shown in the lectures) is also included in the figure. Please inspect the schematic carefully and choose which of the following statements is true: I. Injection of the white experimental transgene into the vermillion mutant embryo will not change the vermillion mutant phenotype II. Injection of the white experimental transgene in the rosy mutant embryo will change rosy eye colour to red (wild type) III. Injection of the white experimental transgene in the white mutant embryo will not change the white mutant phenotype IV. Injection of the white experimental transgene in the rosy mutant…
The transcription factor Pax6 is required continually during the life of a mouse (or a human) for the development of the retina. Homozygous Pax6 knockout mice die soon after birth because Pax6 protein is also required in essential organs, such as the pancreas. a) In order to study the role of Pax6 in eye development a researcher wants to generate a mouse that expresses Pax6 everywhere except in its eyes. Describe how you could construct such a mouse by floxing the gene. Is it possible to achieve the same end with a transgene? (Hint: think about using cDNA and RNAI) b) Suppose you want to create a mouse similar to that in part (a), but one where the eye cells from Pax6 function has been removed and now express a gene that specifies a green fluorescent protein (GFP). Marking the cells in this way will allow the investigators to see the shapes of the Pax6- eye cells more easily than if they did not express GFP. Diagram a Pax6 gene construct that would enable the researcher to do this…
A single UASG regulates the expression of threegenes, all of which are adjacent: GAL7 and GAL10 asdescribed in Problem 8, and also GAL1.a. Would you expect these genes to be transcribedinto individual transcripts, or to be cotranscribed asone mRNA? Explain.b. How could you determine experimentallywhether each gene is transcribed separately orinstead that the three are cotranscribed into asingle mRNA?c. GAL1 and GAL10 are not only adjacent to eachother, but also are transcribed divergently with asingle UASG between them. Describe experimentsusing GFP and RFP transgenes that would allowyou to determine which of the four GAL4 bindingsites in this UASG element is (are) important for thetranscription of GAL1 and/or GAL10.
Based on its sequence similarity to Apobec1,which deaminates Cs to Us in RNA, activation-induceddeaminase (AID) was originally proposed to work on RNA.But definitive experiments in E. coli demonstrated thatAID deaminates Cs to Us in DNA. The authors of the paperexpressed AID in bacteria and followed mutations in aselectable gene. They found that AID expression increasedmutations about fivefold above the background level inthe absence of AID expression. More importantly, theyfound that 80% of the induced mutations were G→A orC→T. Does this fit with your expectation if AID-inducedmutations arose by deamination of C to U in the DNA?
The transcription factor Pax6 is required continuallyduring the life of a mouse (or a human) for the development and maintenance of the retina. HomozygousPax6 knockout mice die soon after birth because Pax6protein is also required in essential organs, such as thepancreas.a. In order to study the role of Pax6 in eye development, a researcher wants to generate a mouse thatexpresses Pax6 everywhere except in its eyes.Describe how you could construct such a mouse.b. Suppose the scientist wants to create a mousesimilar to that in part (a), but in which the eye cellsfrom which Pax6 function has been removed nowexpress GFP. Marking the cells in this way willallow the investigator to see the shapes of the Pax6−eye cells more easily than if they did not expressGFP. Diagram a Pax6 gene construct that wouldenable her to do this experiment.