P2 Discussion

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Revise on Central Dogma

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http://academic.pgcc.edu/~kroberts/Lecture/Chapter%207/dogma.html

Universal Genetic Code (translation)

http://plato.stanford.edu/entries/information-biological/GeneticCode.png

Name 3 Letter 1 Letter Alanine Ala A Cysteine Cys C Aspartic Acid Asp D Glutamic Acid Glu E Phenylalanine Phe F Glycine Gly G Histidine His H Isoleucine Ile I Lysine Lys K Leucine Leu L Methionine Met M Asparagine Asn N Proline Pro P Glutamine Gln Q Arginine Arg R Serine Ser S Threonine Thr T Valine Val V Tyrosine Tyr YTryptophan Trp W

64 genetic codes 20 amino acids

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Physiochemical properties:http://instruct.westvalley.edu/svensson/Cells/07molecules.htm

• R: variable side chain: strongly influences the property of amino acids.

• Each amino acid: different chemical properties (large vs small, hydrophobic vs hydrophilic)

Discussion: six-frame translation

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Emboss six-pack: in-silico translate p53 mRNA sequence in 6-frames

In-vivo Translation:Begins with start codon & ends at stop codonIntrons & UTRs are not translated (spliced)

In-silico Translation:6 reading frames (3 forward & 3 reverse strand)Look for stop codons for each reading frameOutput translations from all open reading frames enclosed by stop codonsTranslate entire sequence including introns & UTRs

Open Reading Frame (ORF)

• A fragment of DNA sequence that has the potential to encode a peptide or protein

• A stretch of triplet codons that encode amino acids with an initiator codon (ATG) at one end and a stop codon (TAA, TAG or TGA) at the other

• Computational translation (in-silico) (e.g., six-pack) Output ORFs: enclosed by stop codons Not necessarily begin with a start codon Translate according to the 6-reading frames Theoretical prediction

ACU UAG CUA CGA UCG ACA UCG CUG U Reading Frame 1

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In-silico 6-frame Translation5’-ACAGCGATGTCGATCGTAGCTAAGT-3’ Forward DNA

3’-UGUCGCUACAGCUAGCAUCGAUUCA-5’ mRNA1(forward DNA)5’-ACAGCGAUGUCGAUCGUAGCUAAGU-3’ mRNA2(complement DNA)

Transcription

5’-ACUUAGCUACGAUCGACAUCGCUGU-3’ mRNA1(forward DNA)

Translation

A CUU AGC UAC GAU CGA CAU CGC UGU Reading Frame 2 AC UUA GCU ACG AUC GAC AUC GCU GU Reading Frame 3

Frame1: CDAKLQMLDSGRHKS*KSLDFDFKHY*GTWQVN

In-silico ORF1: CDAKLQMLDSGRHKSIn-silico ORF2: KSLDFDFKHYIn-silico ORF3: GTWQVN

In-vivo Translation: start M, end at first *

3’-TGTCGCTACAGCTAGCATCGATTCA-5’ Complement

Reverse strand: Reading Frame 4,5,6

Naming of Frames – Caution!

• There is still no standard (different groups call them differently)– 1, 2, 3, 4, 5, 6

– +1, +2, +3, -1, -2, -3

– 1, 2, 3, 1’, 2’, 3’

• Be careful when cross-referencing frame names between two or more programs– know which frames the numbers are really referring to

Translation tool: Wemboss sixpack• Outseq: lists all the ORFs obtained from the six translation

frames.

• Outfile: shows users the input sequence, its complementary strand, and all the open reading frames (ORFs) in the six translation frames