DNA Read, Write and Edit

Part 1: Benchling & In-silico Gel Art

Part 2: Gel Art - Restriction Digests and Gel Electrophoresis

No access to lab

Part 3: DNA Design Challenge

3.1. LUCIFERASE

Bioluminescent art is based on the organic production of visible light by living organisms. This light is produced through the oxidation of luciferin, which is catalyzed by an enzyme called luciferase. For this week’s homework, we will focus on the protein coding for this enzyme, which was first identified in the firefly species (Photinus Pyrlis) [1].

[1] De Wet J.R. et al. Firefly luciferase gene: structure and expression in mammalian cells. Mol Cell Biol (1987). https://pmc.ncbi.nlm.nih.gov/articles/PMC365129/

Sources

NCBI database search: https://www.ncbi.nlm.nih.gov/protein/BAF48396.1

Uniprot database search: https://www.uniprot.org/uniprotkb/P08659/entry#sequences

Protein Sequence

550 amino acids

1 medaknikkg papfypledg tageqlhkam kryalvpgti aftdahievn ityaeyfems

61 vrlaeamkry glntnhrivv csenslqffm pvlgalfigv avapandiyn erellnsmni

121 sqptvvfvsk kglqkilnvq kklpiiqkii imdsktdyqg fqsmytfvts hlppgfneyd

181 fvpesfdrdk tialimnssg stglpkgval phrtacvrfs hardpifgnq iipdtailsv

241 vpfhhgfgmf ttlgylicgf rvvlmyrfee elflrslqdy kiqsallvpt lfsffakstl

301 idkydlsnlh eiasggapls kevgeavakr fhlpgirqgy gltettsail itpegddkpg

361 avgkvvpffe akvvdldtgk tlgvnqrgel cvrgpmimsg yvnnpeatna lidkdgwlhs

421 gdlaywdede hffivgrlks likykgyqva paelesillq hpnifdagva glpdddagel

481 paavvvlehg ktmtekeivd yvasqvttak klrggvvfvd evpkgltgkr darkireili

541 kakkggkskl

3.2. Reverse Translate: Protein sequence to DNA sequence

Source

NCBI database search for P.pyralis (firefly) luciferase gene: https://www.ncbi.nlm.nih.gov/nuccore/M15077

Luciferase protein DNA sequence

    1 ctgcagaaat aactaggtac taagcccgtt tgtgaaaagt ggccaaaccc ataaatttgg
   61 caattacaat aaagaagcta aaattgtggt caaactcaca aacattttta ttatatacat
  121 tttagtagct gatgcttata aaagcaatat ttaaatcgta aacaacaaat aaaataaaat
  181 ttaaacgatg tgattaagag ccaaaggtcc tctagaaaaa ggtatttaag caacggaatt
  241 cctttgtgtt acattcttga atgtcgctcg cagtgacatt agcattccgg tactgttggt
  301 aaaatggaag acgccaaaaa cataaagaaa ggcccggcgc cattctatcc tctagaggat
  361 ggaaccgctg gagagcaact gcataaggct atgaagagat acgccctggt tcctggaaca
  421 attgcttttg tgagtatttc tgtctgattt ctttcgagtt aacgaaatgt tcttatgttt
  481 ctttagacag atgcacatat cgaggtgaac atcacgtacg cggaatactt cgaaatgtcc
  541 gttcggttgg cagaagctat gaaacgatat gggctgaata caaatcacag aatcgtcgta
  601 tgcagtgaaa actctcttca attctttatg ccggtgttgg gcgcgttatt tatcggagtt
  661 gcagttgcgc ccgcgaacga catttataat gaacgtaagc accctcgcca tcagaccaaa
  721 gggaatgacg tatttaattt ttaaggtgaa ttgctcaaca gtatgaacat ttcgcagcct
  781 accgtagtgt ttgtttccaa aaaggggttg caaaaaattt tgaacgtgca aaaaaaatta
  841 ccaataatcc agaaaattat tatcatggat tctaaaacgg attaccaggg atttcagtcg
  901 atgtacacgt tcgtcacatc tcatctacct cccggtttta atgaatacga ttttgtacca
  961 gagtcctttg atcgtgacaa aacaattgca ctgataatga attcctctgg atctactggg
 1021 ttacctaagg gtgtggccct tccgcataga actgcctgcg tcagattctc gcatgccagg
 1081 tatgtcgtat aacaagagat taagtaatgt tgctacacac attgtagaga tcctattttt
 1141 ggcaatcaaa tcattccgga tactgcgatt ttaagtgttg ttccattcca tcacggtttt
 1201 ggaatgttta ctacactcgg atatttgata tgtggatttc gagtcgtctt aatgtataga
 1261 tttgaagaag agctgttttt acgatccctt caggattaca aaattcaaag tgcgttgcta
 1321 gtaccaaccc tattttcatt cttcgccaaa agcactctga ttgacaaata cgatttatct
 1381 aatttacacg aaattgcttc tgggggcgca cctctttcga aagaagtcgg ggaagcggtt
 1441 gcaaaacggt gagttaagcg cattgctagt atttcaaggc tctaaaacgg cgcgtagctt
 1501 ccatcttcca gggatacgac aaggatatgg gctcactgag actacatcag ctattctgat
 1561 tacacccgag ggggatgata aaccgggcgc ggtcggtaaa gttgttccat tttttgaagc
 1621 gaaggttgtg gatctggata ccgggaaaac gctgggcgtt aatcagagag gcgaattatg
 1681 tgtcagagga cctatgatta tgtccggtta tgtaaacaat ccggaagcga ccaacgcctt
 1741 gattgacaag gatggatggc tacattctgg agacatagct tactgggacg aagacgaaca
 1801 cttcttcata gttgaccgct tgaagtcttt aattaaatac aaaggatatc aggtaatgaa
 1861 gatttttaca tgcacacacg ctacaatacc tgtaggtggc ccccgctgaa ttggaatcga
 1921 tattgttaca acaccccaac atcttcgacg cgggcgtggc aggtcttccc gacgatgacg
 1981 ccggtgaact tcccgccgcc gttgttgttt tggagcacgg aaagacgatg acggaaaaag
 2041 agatcgtgga ttacgtcgcc agtaaatgaa ttcgttttac gttactcgta ctacaattct
 2101 tttcataggt caagtaacaa ccgcgaaaaa gttgcgcgga ggagttgtgt ttgtggacga
 2161 agtaccgaaa ggtcttaccg gaaaactcga cgcaagaaaa atcagagaga tcctcataaa
 2221 ggccaagaag ggcggaaagt ccaaattgta aaatgtaact gtattcagcg atgacgaaat
 2281 tcttagctat tgtaatatta tatgcaaatt gatgaatggt aattttgtaa ttgtgggtca
 2341 ctgtactatt ttaacgaata ataaaatcag gtataggtaa ctaaaaa

3.3. Codon optimization

According to the genetic code, there are fewer amino acids than codon possibilities (see chart below, image credit cdn.prod.website-files.com). Thus, in theory, multiple codons can encode for the same amino acid. But in practice, spatial configuration and kinetics factors affect the translation process. For instance, the use of some codons ressembling the STOP codons can interrupt prematurely the translation process. Thus, codon optimization is an important step when designing a nucleotide sequence.

Luciferas protein DNA sequence with Codon-Optimization

(In Progress)