Question

1) How many amino acids long is the protein with the following sequence?

5’ tctaaaatgtcagatgtaaagtgcacatcagtagtcttactctcagttttgtaacaactc 3’

3’ agattttacagtctacatttcacgtgtagtcatcagaatgagagtcaaaacattgttgag 5’

2) Design a set of 6 base pair long primers that scientists could use to amplify the entire sequence of the gene in Q1

3) T/F. A base pair substitution (ie. a point mutation) that introduces a nonsense mutation could be introduced to the sequence encoding the protein in Q1.

Answer 1

1. The protein is produced from the coding strand that is the 3' ---- 5' strand.

So,

Coding DNA strand sequence:

3' - AGA TTT TAC AGT CTA CAT TTC ACG TGT AGT CAT CAG AAT GAG AGT CAA AAC ATT GTT GAG - 5'

mRNA Strand sequence:

5' - UCU AAA AUG UCA GAU GUA AAG UGC ACA UCA GUA GUC UUA CUC UCA GUU UUG UAA CAA CUC - 3'

Aminoacid sequnce:

- Ser-Lys-Met-Ser-Asp-Val-Lys-Cys-Thr-Ser-Val-Val-Leu-Leu-Ser-Val-Leu-STOP -

This protein produced is 18 aminoacids long with the following sequence. The codons that are higlighted in the red color are not translated because they appear after the STOP codon because STOP codon terminates the protein synthesis.

2. Forward Primer: 5' - TGTCAG - 3'

Reverse Primer: 5' - GAGTTG - 3'

Forward Primer is designed on the coding strand and Reverse Primer is designed on the template strand. Reverse Primer will be the reverse complement of the actual sequence to avoid hinderences. In primers 3' end is considered to be the active end because the RNA Polymerase will bind at the 3' end of the primer and initiate transcription further. 5' in the primers is considered to be the inactive end.

3. True

A base pair substitution (ie. a point mutation) that introduces a nonsense mutation could be introduced to the sequence encoding the protein in Q1.

We can introduce a base pair substitution (a point mutation) into the sequence encoding protein in Q1 at 29th position, we are substituting G with T which is highlighted in blue color then that codes UAA which is also highlighted in blue color in the mRNA sequence. UAA codes for STOP codon whereas, previously it was UCA which codes for Ser. This substituion is taking place at the 10th position which reaches the termination abruptly after translating 9 amino acids. Non sense mutation is a point mutation where one base pair substitution leads to shortened or abrupt termination of the protein synthesis.

Coding DNA strand sequence with induced point mutation:

3' - AGA TTT TAC AGT CTA CAT TTC ACG TGT ATT CAT CAG AAT GAG AGT CAA AAC ATT GTT GAG - 5'

mRNA Strand sequence with induced point mutation:

5' - UCU AAA AUG UCA GAU GUA AAG UGC ACA UAA GUA GUC UUA CUC UCA GUU UUG UAA CAA CUC - 3'

Aminoacid sequnce with induced point mutation:

- Ser-Lys-Met-Ser-Asp-Val-Lys-Cys-Thr-STOP -