LOCUS       sequence-488751-        4991 bp    DNA     circular SYN 08-OCT-2025
DEFINITION  Encodes ERK SPARK, a ERK activity reporter, as a Type 3 part to be 
            used in the MTK system.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4991)
  AUTHORS   Fonseca JP, Bonny AR, Kumar GR, Ng AH, Town J, Wu QC, Aslankoohi E, 
            Chen SY, Dods G, Harrigan P, Osimiri LC, Kistler AL, El-Samad H
  TITLE     A Toolkit for Rapid Modular Construction of Biological Circuits in 
            Mammalian Cells.
  JOURNAL   ACS Synth Biol. 2019 Nov 15;8(11):2593-2606. doi: 
            10.1021/acssynbio.9b00322. Epub 2019 Nov 5.
  PUBMED    31686495
REFERENCE   2  (bases 1 to 4991)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 8, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1021/acssynbio.9b00322"; journalName: "ACS Synth Biol"; date: 
            "2019-11-15- 15"; volume: "8"; issue: "11"; pages: "2593-2606"
COMMENT     Sequence Label: MTK3_030
FEATURES             Location/Qualifiers
     source          1..4991
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             join(279..281,282..284,285..995)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             1191..1244
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     regulatory      1251..1260
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1461..1463,1464..1466,1467..2177)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     terminator      complement(2395..2429)
                     /label=lambda t0 terminator
                     /note="minimal transcription terminator from phage lambda 
                     (Scholtissek and Grosse, 1987)"
     CDS             complement(2500..3159)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     3074..3093
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     rep_origin      3341..3929
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3830..3849
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(4415..4449)
                     /label=lambda t0 terminator
                     /note="minimal transcription terminator from phage lambda 
                     (Scholtissek and Grosse, 1987)"
     gap             4529..4628
                     /estimated_length=100
ORIGIN
        1 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
       61 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtagtc
      121 ggtctcatat ggctgaccaa ctgactgaag agtggtcgac ggtaccgcgg accggtcctg
      181 acgtaccacg tacaccagta ggaaaggcaa agctgtcatt ccaattcccg ggcggactcg
      241 agtccggact cagatctggc agcggtggag gcgatatcat ggtgagcaag ggcgaggagc
      301 tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt
      361 tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca
      421 tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg
      481 gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
      541 ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca
      601 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
      661 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca
      721 gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggcc aacttcaaga
      781 tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
      841 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccgccc
      901 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
      961 ccgggatcac tctcggcatg gacgagctgt acaagggatc cggcagtgct ggtggtagtg
     1021 ctggtggtag tgctggtggt agtgctggtg gcagtgctgg tggtagtgct ggtggtagtg
     1081 ctggtggctc tagaggagaa atcgcaaaga gtctgaaaga aattgcaaaa tccttgaagg
     1141 aaatagcgtg gtcactgaaa gaaattgcca agtccctcaa agggggtacc gagggcagag
     1201 gaagtctgct aacatgcggt gacgtcgagg agaatcctgg cccaaagctt gccaccatgg
     1261 cggacgagga gaagctgccg cccggctggg agaagcgcat gagccgcagc tcaggccgag
     1321 tgtactactt caaccacatc actaacgcca gccagtggga gcggcccagc ggcaacagca
     1381 gcagtggtgg caaaaacggg cagggggagc ctgccaggct cgagtccgga ctcagatctg
     1441 gcagcggtgg aggcgatatc atggtgagca agggcgagga gctgttcacc ggggtggtgc
     1501 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
     1561 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
     1621 tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc
     1681 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
     1741 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
     1801 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
     1861 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
     1921 tggccgacaa gcagaagaac ggcatcaagg ccaacttcaa gatccgccac aacatcgagg
     1981 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
     2041 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg
     2101 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca
     2161 tggacgagct gtacaaggga tccggcagtg ctggtggtag tgctggtggt agtgctggtg
     2221 gtagtgctgg tggcagtgct ggtggtagtg ctggtggtag tgctggtggc tctagaacct
     2281 tgagagaaat cgaggaattg ttgaggaaaa tcattgaaga cagcgtgcgc agtgtggccg
     2341 agctggagga cattgagaag tggctgaaga aaatcggatc ctgagaccag accaataaaa
     2401 aacgcccggc ggcaaccgag cgttctgaac aaatccagat ggagttctga ggtcattact
     2461 ggatctatca acaggagtcc aagcgagctc gatatcaaat tacgccccgc cctgccactc
     2521 atcgcagtac tgttgtaatt cattaagcat tctgccgaca tggaagccat cacaaacggc
     2581 atgatgaacc tgaatcgcca gcggcatcag caccttgtcg ccttgcgtat aatatttgcc
     2641 catggtgaaa acgggggcga agaagttgtc catattggcc acgtttaaat caaaactggt
     2701 gaaactcacc cagggattgg ctgaaacgaa aaacatattc tcaataaacc ctttagggaa
     2761 ataggccagg ttttcaccgt aacacgccac atcttgcgaa tatatgtgta gaaactgccg
     2821 gaaatcgtcg tggtattcac tccagagcga tgaaaacgtt tcagtttgct catggaaaac
     2881 ggtgtaacaa gggtgaacac tatcccatat caccagctca ccgtctttca ttgccatacg
     2941 aaattccgga tgagcattca tcaggcgggc aagaatgtga ataaaggccg gataaaactt
     3001 gtgcttattt ttctttacgg tctttaaaaa ggccgtaata tccagctgaa cggtctggtt
     3061 ataggtacat tgagcaactg actgaaatgc ctcaaaatgt tctttacgat gccattggga
     3121 tatatcaacg gtggtatatc cagtgatttt tttctccatt ttagcttcct tagctcctga
     3181 aaatctcgat aactcaaaaa atacgcccgg tagtgatctt atttcattat ggtgaaagtt
     3241 ggaacctctt acgtgcccga tcaatcatga ccaaaatccc ttaacgtgag ttttcgttcc
     3301 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     3361 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     3421 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     3481 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     3541 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     3601 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     3661 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     3721 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     3781 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     3841 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     3901 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     3961 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     4021 ataaccgtag tcggtctcac cctgaattcg catctagatg gtagagccac aaacagccgg
     4081 tacaagcaac gatctccagg accatctgaa tcatgcgcgg atgacacgaa ctcacgacgg
     4141 cgatcacaga cattaaccca cagtacagac actgcgacaa cgtggcaatt cgtcgcaata
     4201 ccgtctcact gaactggccg ataattgcag acgaggagca tcgcccttcc ccggccctca
     4261 ggtaagagga ccaaataccg tagccgtttc caatttcagt cctttagcgc cacctggtgc
     4321 taactactct atcacgcttt tatccaataa ctacctttgt aaatttcctt tcaaaagttc
     4381 tggccgggcg cggtggcaac gtgagaccag accaataaaa aacgcccggc ggcaaccgag
     4441 cgttctgaac aaatccagat ggagttctga ggtcattact ggatctatca acaggagtcc
     4501 aagcgagctc gatatcaaat tacgccccnn nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn
     4561 nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn
     4621 nnnnnnnngg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     4681 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     4741 ccgtagtcgg tctcatatgg ctgaccaact gactgaagag tggtcgacgg taccgcggac
     4801 cggtcctgac gtaccacgta caccagtagg aaaggcaaag ctgtcattcc aattcccggg
     4861 cggactcgag tccggactca gatctggcag cggtggaggc gatatcatgg tgagcaaggg
     4921 cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg
     4981 ccacaagttc a
//
