BOC314 PRACTICAL 3
TRANSLATION OF A NUCLEOTIDE SEQUENCE CONTAINING INTRONS
A.
RETRIEVAL OF SEQUENCE
1.
Use Entrez to retrieve the nucleotide sequence of Rhodosporidium paludigenum epoxide
hydrolase gene”.
2.
Copy all the information to a Microsoft Word document. (Remember to adjust the Margins, Font
and Font size as in Practical 2). Insert a page break at the bottom of the page.
B.
TRANSLATION OF NUCLEOTIDE SEQUENCE
1.
Go to the following site: http://tw.expasy.org/. Click on ‘full list’ under “Tools and Software
packages”, click on Translate under “DNA -> Protein”.
2.
Paste the nucleotide sequence into the box.
3.
Select the Output format Verbose (“Met”, “Stop”, spaces etc.)
4.
Translate the sequence.
5.
Copy and paste the Results of Translation in your word file (all six reading frames) and insert a
page break at the bottom of the page. Save the file. (Note that all the reading frames contain a
number of ‘STOP’ codons).
C.
1.
REMOVAL OF INTRONS FROM NUCLEOTIDE SEQUENCE OF EPH1 GENE
Copy only the nucleotide sequence of the Rhodosporidium paludigenum epoxide hydrolase
gene to a new page (below the translated sequence).
2.
Use this copy and the information of the gene (which you retrieved from Entrez) to physically
remove all the introns of the EPH1 gene.

First locate the start and stop codon, change the font colour to blue and bold these
sections of the region before the start and after the stop codon.

Secondly, select each intron (using the information provided from the sequence report
retrieved in A) and change the font colour to red (and bold) of each intron.

When all the introns have been labelled like this COPY THIS TO A NEW PAGE, use this 2nd
copy to delete all the introns (in red font) as well as the non-coding regions before and
after the coding region (blue font).
Do not delete anything else (e.g. the baspair
numbering)
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3.
When all the introns have been removed, insert a page break at the bottom of the page. Copy the
resulting sequence (without the introns) in the box of the Translation Tool and translate the
sequence as above.
5.
Copy and paste the Results of Translation in file Practical3.doc (this time only copy the correct
translated reading frame to your report). You can identify the correct reading frame by comparing
all the reading frames with the protein sequence retrieved in A.
6.
Copy the reading frame you selected above and change all the MET (for methionine) to M and
delete the STOP before the next step.
D. MULTIPLE ALIGNMENTS OF SEQUENCES
1.
Perform a multiple alignment with the protein sequence created by the translation tool and the
protein sequence given in the sequence report you obtained in section A. (http://www2.ebi.ac.uk)
(Toolbox: ClustalW)
2.
Copy the results of the alignment to the bottom of your document. SAVE the file.
3.
The two sequences must align 100%.
E.
REPORT

Part 1: Nucleotide sequence of Rhodosporidium paludigenum epoxide hydrolase gene.

Part 2: Nucleotide sequence translation.

Part 3: Nucleotide sequence of the Rhodosporidium paludigenum epoxide hydrolase with
indicated introns.

Part 4: Nucleotide sequence of the Rhodosporidium paludigenum epoxide hydrolase with
removed introns.

Part 4: Multiple alignment of two protein sequences

Total marks = 27
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