You are interested in studying eukaryotic transcription factors and have develop

Question

You are interested in studying eukaryotic transcription factors and have developed an in vitro transcription system using a segment of DNA that includes a transcribed gene and its predicted upstream regulatory region (see figure below). The transcription of this DNA occurs in a defined system in which you add purified RNA polymerase II and basal factors You perform a series of experiments that compare the efficiency of transcription in your defined system with the efficiency of transcription in a crude nuclear extract. You test the two systems with your WT DNA and various deletion mutants that you have generated (see figure below). The results of your study are shown in the table: a) Why is there no transcription from the -11 deletion template? b) Why do the results for the WT DNA differ between the nuclear extract and the defined system? c) Compare the results obtained for the deletion mutants when tested in the nuclear extract and the defined system. Propose a model to explain the differences - be as specific as you can.

Explanation / Answer

a] yes there will be no trancription from the -11 deletion of template from the figure we can that there is a deletion in the promoter region TATA BOX so , there is no trancription

b] nuclear extarcts has the meachinary where it can transcribe it happens independenly , as the nuclear has the enzymes , proteins and transcrptional factors so it need not depend on the external factors ,so the transcrption process is faster in the nuclear extarct than in a defined system

c] efficency of trnacritption can be studied using the nuclar system and defined system can be done with the deletion of 5 prime flanking dna region   in the nuclaer systen deltionof dna in this region causes the expression of the gene but in defined system ther will no transcription , you can also study with deletion mutation with the help of cloning the dna and see for the expression of gene

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