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- For each of the E. coli strains that follow, indicate theeffect of the genotype on the expression of the trpEand trpC genes in the presence or absence of tryptophan. [In the wild type (R+ P+ o+ att+ trpE+ trpC+),trpC and trpE are fully repressed in the presence oftryptophan and are fully expressed in the absence oftryptophan.]R = repressor gene; Rnproduct cannot bind tryptophan; R− product cannot bind operatoro = operator for the trp operon; o− cannot bind repressoratt = attenuator; att− is a deletion of the attenuatorP = promoter; P− is a deletion of the trp operonpromotertrpE− and trpC− are null (loss-of-function) mutationsa. R+ P− o+ att+ trpE+ trpC+b. R− P+ o+ att+ trpE+ trpC+c. RnP+ o+ att+ trpE+ trpC+d. R− P+ o+ att− trpE+ trpC+e. R+ P+ o− att+ trpE+ trpC−/R− P+ o+ att+trpE− trpC+f. R+ P− o+ att+ trpE+ trpC−/R− P+ o+ att+trpE− trpC+g. R+ P+ o− att− trpE+ trpC−/R− P+ o− att+trpE− trpC+Suppose you have just determined the DNA base sequence foran especially strong promoter in Escherichia coli and you areinterested in incorporating this sequence into an expressionvector. Describe the steps you would use. What precautionsare necessary to be sure that this promoter actually works asexpected in its new location?CTP synthetase catalyzes the glutamine-dependent conversion of UTP to CTP. The enzyme is allosterically inhibited by the product, CTP. Mamma- lian cells defective in this allosteric inhibition are found to have a complex phenotype: They require thymidine in the growth medium, they have unbal- anced nucleotide pools, and they have an elevated spontaneous mutation rate. Explain the likely basis for these observations.
- Another mutant for of the Lac promoter demonstrates the characteristics below compared to the wild-type promoter. The RNA Polymerase Holoenzyme binds tighter to the mutant promoter, but has reduced promoter activity in an assay with the promoter fuse to the gene for Green Fluorescent Protein (GFP). Provide a plausible explanation for this data. Lac Promoter Activity punog Binding of RNA Pol Holoenzymes to Promoters 120 100 80 60 40 20 0 0 20 40 Lac Promoter X 60 [DNA] nM 80 100 120 GFP Fluorecence 100 90 80 70 60 50 40 30 20 10 0 Lac Promoter XConsider a mutant form of the lac repressor protein whose ΔGo for binding DNA differs from that of the wild-type protein by 2 kcal/mol, the approximate free energy equivalent of a single hydrogen bond. What is the difference (i.e., the ratio) of the Kd values (for DNA binding) of the wild-type versus the mutant lac repressor protein?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]
- explain please: Spike-Driven Syncytia Formation Is Coupled to S20 Fragment Generation in the Presence of ACE2. To investigate the functional and biochemical signatures of spike (S) protein upon engaging its receptor ACE2 in live cells, we first utilized a cell–cell fusion assay to obtain potentially cleaved spike protein products from syncytia . In this system, HEK293T cells transfected with plasmids encoding WT spike readily formed syncytia after coculturing with HEK293T cells expressing human and Vero E6-ACE2, as well as colorectal adenocarcinoma Caco-2 and lung adenocarcinoma Calu-3 cells expressing endogenous ACE2. We then collected these adherent syncytia and immunoblotted for cleaved spike species using a rabbit polyclonal antibody specifically detecting the S2 ectodomain through standard reducing Tris-glycine sodium dodecyl sulfate polyacrylamide gel electrophoresis. This experiment revealed that spike expression in HEK293T cells displayed full-length S and autocleaved S2…In addition to Tc1, the C. elegans genome contains otherfamilies of DNA transposons such as Tc2, Tc3, Tc4, andTc5. Like Tc1, their transposition is repressed in thegerm line but not in somatic cells. Predict the behaviorof these elements in the mutant strains where Tc1 is nolonger repressed due to mutations in the RNAi pathway.Justify your answer.In the galactose operon of E. coli, a repressor, encoded by the galR gene, binds to an operator site,galo, to regulate the expression of three structuralgenes, galE, galT, and galK. Expression is inducedby the presence of galactose in the media. For eachof the strains listed, would the cell show constitutive, inducible, or no expression of each of the structural genes? (Assume that galR− is a loss-of-functionmutation.)a. galR−galo+galE+galT+galK+b. galR+galocgalE+galT+galK+ c. galR−galo+galE+galT+galK−/galR+galo+galE−galT+galK+d. galR−galocgalE+galT+galK−/galR+galo+galE−galT+galK+
- Given the following Western blot results showing levels of proteins over time in three yeast cell extracts after the induction of high level expression of the CDH1 protein. WT GAL-CDH1 GAL-CDH1, cdc23-1 0 30 60 120 0 30 60 120 0 30 60 120 Time (min) - Pds1 - Clb2 - Ase1 - Kar2 Note: The GAL-CDH1 indicates that the yeast strain contains a transgenic cdh1 gene under the control of the GAL promoter. This means that the researcher can control expression of that cdh1 gene. In these three experiments (three yeast strains), time 0 is the point at which GAL- controlled genes begin to get expressed (WT lacks transgenes). In this experiment, four proteins are being probed for in three different yeast strains (1. wild-type, 2. a strain containing the cdh1 gene under GAL control (see note), and 3. the GAL-CDH1 strain that is also harboring a cdc23 loss of function allele (allele 1), which is a component of the A The upper (larger) band in the Ase1 lanes is non-specific background band; disregard…Negative supercoiling of DNA favors the transcription of genes because it facilitates unwinding. However, not all promoter sites are stimulated by negative supercoiling. The promoter site for topoisomerase II itself is a noteworthy exception. Negative supercoiling decreases the rate of transcription of this gene. Propose a possible mechanism for this effect and suggest a reason why it may occur.Given the following Western blot results showing levels of proteins over time in three yeast cell extracts after the induction of high level expression of the CDH1 protein. WT GAL-CDH1 GAL-CDH1, cdc23-1 0 30 60 120 0 30 60 120 0 30 60 120 Time (min) | Pds1 - Clb2 - Ase1 - Kar2 Note: The GAL-CDH1 indicates that the yeast strain contains a transgenic cdh1 gene under the control of the GAL promoter. This means that the researcher can control expression of that cdh1 gene. In these three experiments (three yeast strains), time O is the point at which GAL- controlled genes begin to get expressed (WT lacks transgenes). which is a component of the A In this experiment, four proteins are being probed for in three different yeast strains (1. wild-type, 2. a strain containing the cdh1 gene under GAL control (see note), and 3. the GAL-CDH1 strain that is also harboring a cdc23 loss of function allele (allele The upper (larger) band in the Ase1 lanes is non-specific background band; disregard it.…