scription Factors (35 points) Trans ted in how transcription factors TF1, TF2, a
ID: 278199 • Letter: S
Question
scription Factors (35 points) Trans ted in how transcription factors TF1, TF2, and TF3 activate the LOK gene. You 3) designed and synthesized a reporter construct that contains the promoter of the LOK gene followed by a reporter gene (shown below) construct and different combinations of TF1, TF2, and TF3. ). You then perform a gel shift assay and a reporter activity assay with this reporter Gel shift A) Based on the gel shift assay, explain the LOK Promoter DNA binding behavior of the three transcription factors and their combinations. (40 words) Reporter s100 80 60 40 B) Based on the reporter assay, explain the ability of the three transcription factors and their combinations to activate transcription of the LOK gene. (30 words) DNA TF1 TF2 TF3--++ C) Based on A) and B) draw a model of how the reporter gene. the three transcription factors bind the LOK promoter and activate Reporter Gene LOK PromoterExplanation / Answer
Answer (A)
The protein- DNA interation is the central control of the cellular processes like transciption, etc. Trancription Factors are protein which play major role in the DNA trancription, when the TFs interact with promoter , it activates trancription. The binding of TF to DNA increases the size of DNA and the band in gel appear on upper side. Here Lane 1 only DNA, Lane 2 and TF1 is binding to DNA so band is corresponds to 4, Lane 3 and 4 no binding in to the TF2 and TF3. In the presence of TF1 , TF2 is showing interaction with DNA in Lane 5, but TF3 is not in Lane6. We can conclude that TF1 is directly interacting to DNA and favours the binding of TF2, the binding TF2 favours the interaction of TF3 to DNA. The bottom line is all three transcription factors are required for the transcription.
Answer (B)
The expression of reporter gene confirms the transcription to happen, here in the presence of TF1 and TF2 combination there is activity of reporter gene is seen upto 50 %. the combination of TF1 and TF3 alone shows no interaction, it means TF3 disfavour the binding of TF1 to DNA.The highest activity is seen in combination of all TFs, so all trancription factors are important for efficient expression.
Answer (C)
binding sequence-
LOK promotor sequence in DNA+TF1+TF2+TF3
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