Answer the Questions
INVESTIGATION 1: IDENTIFY THE TRANSCRIPTION UNIT
Q10. Which base position(s) would you assign as the TSS of the tra gene based on the available evidence? Describe your reasoning.
Q12. Are there any perfect matches to the Inr consensus sequence (Figure 11)? What are the coordinates and orientation of these matches? What about the TATA Box motif? Are these signals in good agreement with the beginning of the transcription unit?
Q18. Based on your analysis above, which position is the best choice for the termination signal? Describe your reasoning.
INVESTIGATION: MRNA PROCESSING
Q7.Scroll up to the to the ‘cDNA BT028774’ area. After which coordinate (number in the cDNA) do you see the polyadenylation track (in lower case black letters)?
Q8.How many ‘A’ ribonucleotides have been added to the tra mRNA (represented in the cDNA)?
Q9.Locate the AATAAA termination signal in the cDNA sequence. How many nucleotides 3’ of the final ‘A’ in the signal sequence does the poly(A) run start? (This number should be between 11-30 nucleotides.)
Q10. Which two nucleotides are found just after the end of the first exon of tra-RA? __________ Repeat this determination, identifying the two nucleotides at the start of intron 2 of tra-RA. _______
These two nucleotides are a signal for splicing to occur at the 5’ end of an intron; these represent the first two bases of the intron, often called the donor site (or 5’ splice site).
Q11. At which base does exon 1 end?
Q12. Which two nucleotides are found right before the start of tra-RA exon 2?
Q13. Which two nucleotides are found right before the start of tra-RA exon 3?
Q14. At which base does exon 2 of tra-RA begin? What is its coordinate?
INVESTIGATION 1: EXAMINING RNA-SEQ DATA
Q6. Using the information you’ve gathered so far, make a diagram of the tra-RA (female specific) isoform with 3 exons and 2 introns. Represent exons as rectangular boxes and introns as lines connecting the boxes. Number each exon and intron (start from the left with ‘exon 1’).
Q17. Using the information you’ve gathered so far, make a graphical picture of the traRA (female specific) isoform with 3 exons and 2 introns. Number each exon and intron at the corresponding DNA coordinates. Add the coordinates for first and last nucleotide of the exons that you have found so far. Add the sequences of the splice donor and splice acceptor sites at the appropriate locations.
MODULE 4 HOMEWORK: SPLICING
Q11. Using the information you’ve gathered so far, make a graphical picture of the spd2 gene with 3 exons and 2 introns. Number each exon and intron. Add the coordinates for first and last nucleotide of the exons that you have found so far. Add the sequences of the splice donor and splice acceptor sites at the appropriate locations. Finally, add a bent arrow for the transcription start site.
INVESTIGATION 1: EXAMINING OPEN READING FRAMES (ORFS) IN THE TRA GENE
Q17. Let’s consolidate all the data we found above in one place:
Gene model for tra-RA
Coordinate for start of translation: __________________
Coordinate for last base of exon 1: ____________________
Coordinate for first base of exon 2: ___________________
Coordinate for last base of exon 2: ____________________
Coordinate for first base of exon 3: ____________________
Stop codon coordinates: _____________________________
Take the coordinate information above to draw a map of tra-RA using rectangles to represent exons and connecting lines to represent introns. Label the ends of the exons with the appropriate coordinates and indicate the transcription start site for the tra-RA initial transcript. Below this map, provide a map of the processed mRNA after intron removal. Below this map, indicate the regions that are translated into a protein. Give precise coordinates. Color coding may be helpful.
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