Application to find Eukaryotic Open reading frames.

1. Application to find Eukaryotic Open reading frames. Lab

2. Introduction • Finding ORF in Prokaryotic DNA • Finding cds in non-intronic Eukaryotes “genes” • Use perl scripts (from assignment or elsewhere). • Finding ORF in Intronic eukaryotic genes • Compare and contrast results of your program with actual results (gene bank or EBI records) • Test an online gene finding

3. Using your assignment code • Open the file: ORF pal gene.fasta • Find all open reading frames. (This time you must modify your code to translate each codon you can utilise the convertor_hashtable.txt • Compare to file: pal protein sequence.fasta. • Visual inspect the files for start and stops; ORFs.; in which reading frame is the TRUE ORF. • Compare your results to the results in the gene bank record. Annotation and Analysis site: Pal gene • What conclusion can you draw. • Note that the file • ORF pal gene.fasta (has extra bases on either side of the ORF take this into account in your analysis)

4. Predictive translation effect Exons/intron length • Consider the following: • We have the mRNA CDS of 60 bp in length (start…stop) • Let us assume that the intron1 is: • at the end of codon three (position 9) • the length of the intron is 30bp. • Intron 2 occurs at: • the end of codon 10 (position 30) • and is 45 bp in length • What is the effect of the translations: on Exon A and Exon B? Exon A Exon B TAA ATG DNA Strand

5. Predictive translation effect Exons/intron length • Consider the following: • We have the mRNA CDS of 60 bp in length (start…stop) • Let us assume that the intron1 is: • at the end of codon three (position 9) • the length of the intron is 30bp. • Intron 2 occurs at: • at position 29 (at the 3rdbp of codon 10) • and is 45 bp in length • What is the effect of the translations: on Exon A and Exon B? Exon A Exon B TAA ATG DNA Strand

6. Predictive translation effect Exons/intron length • Consider the following: • We have the mRNA CDS of 60 bp in length (start…stop) • Let us assume that the intron1 is: • at the end of codon three (position 9) • the length of the intron is 30bp. • Intron 2 occurs at: • at position 30(the end of codon 10) • and is 43 bp in length • What is the effect of the translations: on Exon A and Exon B? Exon A Exon B TAA ATG DNA Strand

7. Effect of Translation • Example 1 no effect all multiples of 3 • Example 2 the last residue of exon 2 is incorrect. The residue for Exon 3 is correct. (but starts at bp 2 of first codon) • Example 3 last expn is in different reading frame. • Refer to Incorrect_translation_examples.rar

8. Finding start and stop codons and potential ORF • Download the file. (TUBAC3 gene complete sequence) (see last lecture) • Using “your” perl module and perl script determine the location of the start and stop codons. • Write a script that will “attempt” to find at least one ORF. Clearly state how, using your background knowledge of gene expression, you are going to try and resolve this problem. • Using your program: determine the position of an (the) ORF(s) for this gene. • Discuss how your findings compare with the results obtained in the genebank record file . You can use the graphics image

9. Intronic DNA gene • Go to the understanding Bioinformatics student resources and down load the PRISSEL (CDK10 fasta file). It can also be downloaded from the sample files in the lecture on finding Eukaryotic genes. • Run your application to find ORF. Comapre this to the actual results. • Discuss your results (in relation to the known experimental results; you can find the actual results in understanding ioinformatics or refer to slide in the lecture: finding Eukaryotic genes.

10. Online Gene finding program • In the student resources you should find links to various online gene predicting applications. • Use one to analyse the ALDH10 exon 1 and exon 2 of the ALDH10 gene.