LOCUS       sequence-409576-        7872 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Reporter circuit.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7872)
  AUTHORS   Wauford N, Patel A, Tordoff J, Enghuus C, Jin A, Toppen J, Kemp ML, 
            Weiss R
  TITLE     Synthetic symmetry breaking and programmable multicellular structure
            formation.
  JOURNAL   Cell Syst. 2023 Sep 20;14(9):806-818.e5. doi: 
            10.1016/j.cels.2023.08.001. Epub 2023 Sep 8.
  PUBMED    37689062
REFERENCE   2  (bases 1 to 7872)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.cels.2023.08.001"; journalName: "Cell Syst"; date: 
            "2023-09-20- 20"; volume: "14"; issue: "9"; pages: "806-818.e5"
COMMENT     Sequence Label: pJTT002
FEATURES             Location/Qualifiers
     source          1..7872
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(2..26)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(116..135)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    181..236
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(235..254)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     promoter        1210..1421
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     intron          1422..2360
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     2317..2337
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     polyA_signal    3252..3300
                     /note="synthetic polyadenylation signal"
     protein_bind    3391..3424
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Groth 
                     et al., 2000)"
     regulatory      3435..3444
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3441..3443,3444..3446,3447..4160)
                     /codon_start=1
                     /product="enhanced YFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3486..3507)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3747..3766)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4094..4115
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4213..4261
                     /note="synthetic polyadenylation signal"
     regulatory      4412..4421
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4418..5110
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGH"
     polyA_signal    5175..5223
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(5276..5294)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5457..5476)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             5468..6277
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(5535..5554)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      6432..7020
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6921..6940
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    7148..7168
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     protein_bind    7186..7223
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attB site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     protein_bind    7233..7257
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             7273..7872
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(7273..7292)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7769..7789
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
ORIGIN
        1 tacccagctt tcttgtacaa agtggtacgc gtgaattcac tcctcaggtg caggctgcct
       61 atcagaaggt ggtggctggt gtggccaatg ccctggctca caaataccac tgagatcttt
      121 ttccctctgc caaaaattat ggggacatca tgaagcccct tgagcatctg acttctggct
      181 aataaaggaa atttattttc attgcaatag tgtgttggaa ttttttgtgt ctctcactcg
      241 gaaggacata tgggagggca aatcatttaa aacatcagaa tgagtatttg gtttagagtt
      301 tggcaacata tgcccatatg ctggctgcca tgaacaaagg ttggctataa agaggtcatc
      361 agtatatgaa acagccccct gctgtccatt ccttattcca tagaaaagcc ttgacttgag
      421 gttagatttt ttttatattt tgttttgtgt tatttttttc tttaacatcc ctaaaatttt
      481 ccttacatgt tttactagcc agatttttcc tcctctcctg actactccca gtcatagctg
      541 tccctcttct cttatggaga tccctcgacg aggaagcaga gcgtaaaacg acggccaggc
      601 agtaccggag ttctgcaatt catgtgatct gcgcctggga gctcacgggg acagcccccc
      661 cccaaagccc ccagggatgt aattacgtcc ctcccccgct agggggcagc agcgagccgc
      721 ccggggctcc gctccggtcc ggcgctcccc ccgcatcccc gagccggcag cgtgcgggga
      781 cagcccgggc acggggaagg tggcacggga tcgctttcct ctgaacgctt ctcgctgctc
      841 tttgagcctg cagacacctg gggggatacg gggaaaaagc taggctgaaa gaggatctta
      901 gaatgacacg cgcctgggag ctcacgggga cagccccccc ccaaagcccc cagggatgta
      961 attacgtccc tcccccgcta gggggcagca gcgagccgcc cggggctccg ctccggtccg
     1021 gcgctccccc cgcatccccg agccggcagc gtgcggggac agcccgggca cggggaaggt
     1081 ggcacgggat cgctttcctc tgaacgcttc tcgctgctct ttgagcctgc agacacctgg
     1141 ggggatacgg ggaaaaagct ggatcataac tcgacgtgca tactttaatt ggctccggtg
     1201 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgtgg ggaggggtcg
     1261 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
     1321 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
     1381 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
     1441 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
     1501 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
     1561 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
     1621 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
     1681 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
     1741 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
     1801 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
     1861 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
     1921 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     1981 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     2041 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     2101 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     2161 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     2221 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     2281 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     2341 gtttttttct tccatttcag gtggatccca agggcagagg agtagtgccc caactggggt
     2401 aacctttgag ttctctcagt tgggggcgta gggtctagtt ttaaagttgg ttattagtta
     2461 ctgtgatatt tatcacggta cccaataacc aatgaaaggt gccaccatga gcgagctgat
     2521 taaggagaac atgcacatga agctgtacat ggagggcacc gtggacaacc atcacttcaa
     2581 gtgcacatcc gagggcgaag gcaagcccta cgagggcacc cagaccatga gaatcaaggt
     2641 ggtcgagggc ggccctctcc ccttcgcctt cgacatcctg gctactagct tcctctacgg
     2701 cagcaagacc ttcatcaacc acacccaggg catccccgac ttcttcaagc agtccttccc
     2761 tgagggcttc acatgggaga gagtcaccac atacgaagac gggggcgtgc tgaccgctac
     2821 ccaggacacc agcctccagg acggctgcct catctacaac gtcaagatca gaggggtgaa
     2881 cttcacatcc aacggccctg tgatgcagaa gaaaacactc ggctgggagg ccttcaccga
     2941 gacgctgtac cccgctgacg gcggcctgga aggcagaaac gacatggccc tgaagctcgt
     3001 gggcgggagc catctgatcg caaacatcaa gaccacatat agatccaaga aacccgctaa
     3061 gaacctcaag atgcctggcg tctactatgt ggactacaga ctggaaagaa tcaaggaggc
     3121 caacaacgag acatacgtcg agcagcacga ggtggcagtg gccagatact gcgacctccc
     3181 tagcaaactg gggcactgag cttccactgg attgtacaat taccaactga tctagaaccg
     3241 gtcatggccg caataaaata tctttatttt cattacatct gtgtgttggt tttttgtgtg
     3301 ttcgaactag atgctgtcga ccgatgcgct cgcggaggta tcacaggaga atttcagtac
     3361 ttaagtttgt acaaaaaaga gcagatgcgg gtgccagggc gtgcccttgg gctccccggg
     3421 cgcgtactcc aggtgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc
     3481 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
     3541 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
     3601 tgcccgtgcc ctggcccacc ctcgtgacca ccttcggcta cggcctgcag tgcttcgccc
     3661 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
     3721 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
     3781 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
     3841 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
     3901 tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg
     3961 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
     4021 tgctgctgcc cgacaaccac tacctgagct accagtccaa gctgagcaaa gaccccaacg
     4081 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca
     4141 tggacgagct gtacaagtga gcttccactg gattgtacaa ttaccaactg atctagaacc
     4201 ggtcatggcc gcaataaaat atctttattt tcattacatc tgtgtgttgg ttttttgtgt
     4261 gttcgaacta gatgctgtcg accgatgcgc ttacggaggt atcacaggag aatttcagta
     4321 cttaagtttg tacaaaaaag agcagaaagg tagcgtcaac gataggtgta actgtcgtgt
     4381 ttgtaacggt acttccaaca gctggcgagg tgccaccatg gtgagcgagc tgattaagga
     4441 gaacatgcac atgaagctgt acatggaggg caccgtgaac aaccaccact tcaagtgcac
     4501 atccgagggc gaaggcaagc cctacgaggg cacccagacc atgagaatca aggcggtcga
     4561 gggcggccct ctccccttcg ccttcgacat cctggctacc agcttcatgt acggcagcaa
     4621 aaccttcatc aaccacaccc agggcatccc cgacttcttt aagcagtcct tccccgaggg
     4681 cttcacatgg gagagagtca ccacatacga agatgggggc gtgctgaccg ctacccagga
     4741 caccagcctc caggacggct gcctcatcta caacgtcaag atcagagggg tgaacttccc
     4801 atccaacggc cctgtgatgc agaagaaaac actcggctgg gaggcctcca ccgagacact
     4861 gtaccccgct gacggcggcc tggaaggcag agccgacatg gccctgaagc tcgtgggcgg
     4921 gggccacctg atctgcaacc ttaagaccac atacagatcc aagaaacccg ctaagaacct
     4981 caagatgccc ggcgtctact atgtggacag gagactggaa agaatcaagg aggccgacaa
     5041 agagacatac gtcgagcagc acgaggtggc tgtggccaga tactgcgacc tccctagcaa
     5101 actggggcac aaacttaatt gagcttccac tggattgtac aattaccaac tgatctagaa
     5161 ccggtcatgg ccgcaataaa atatctttat tttcattaca tctgtgtgtt ggttttttgt
     5221 gtgttcgaac tagatgctgt cgaccgatgc gctagagttc gcaattcagg aaagggcctc
     5281 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     5341 ggcacttttc ggggaaatgt gctggcccgt gtctcaaaat ctctgatgtt acattgcaca
     5401 agataaaaat atatcatcat gaacaataaa actgtctgct tacataaaca gtaatacaag
     5461 gggtgttatg agccatattc aacgggaaac gtcgaggccg cgattaaatt ccaacatgga
     5521 tgctgattta tatgggtata aatgggctcg cgataatgtc gggcaatcag gtgcgacaat
     5581 ctatcgcttg tatgggaagc ccgatgcgcc agagttgttt ctgaaacatg gcaaaggtag
     5641 cgttgccaat gatgttacag atgagatggt cagactaaac tggctgacgg aatttatgcc
     5701 tcttccgacc atcaagcatt ttatccgtac tcctgatgat gcatggttac tcaccactgc
     5761 gatccccgga aaaacagcat tccaggtatt agaagaatat cctgattcag gtgaaaatat
     5821 tgttgatgcg ctggcagtgt tcctgcgccg gttgcattcg attcctgttt gtaattgtcc
     5881 ttttaacagc gatcgcgtat ttcgtctcgc tcaggcgcaa tcacgaatga ataacggttt
     5941 ggttgatgcg agtgattttg atgacgagcg taatggctgg cctgttgaac aagtctggaa
     6001 agaaatgcat aaacttttgc cattctcacc ggattcagtc gtcactcatg gtgatttctc
     6061 acttgataac cttatttttg acgaggggaa attaataggt tgtattgatg ttggacgagt
     6121 cggaatcgca gaccgatacc aggatcttgc catcctatgg aactgcctcg gtgagttttc
     6181 tccttcatta cagaaacggc tttttcaaaa atatggtatt gataatcctg atatgaataa
     6241 attgcagttt catttgatgc tcgatgagtt tttctaatca gaattggtta attggttgta
     6301 acactggcag agcattacgc tgacttgacg ggacggaaga tcctttttga taatctcatg
     6361 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     6421 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     6481 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     6541 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     6601 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     6661 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     6721 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     6781 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     6841 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     6901 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     6961 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     7021 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     7081 ttctttcctg cgttatcccc tgattctgtg gcatgcgtat cgatgtcgac gttaacgcta
     7141 gtgatatcaa ctttgtatag aaaagttggc tccgaatttc tcgagggctt gtcgacgacg
     7201 gcggtctccg tcgtcaggat catgaattcg acccaagttt gtacaaaaaa gcaggctgaa
     7261 cggcctgcca ccatgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc
     7321 ccccgggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc
     7381 gtcgacccgg accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc
     7441 gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg
     7501 accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc
     7561 gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct ggcgccgcac
     7621 cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga ccaccagggc
     7681 aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg cgccggggtg
     7741 cccgccttcc tggagacctc cgcgccccgc aacctcccct tctacgagcg gctcggcttc
     7801 accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag
     7861 cccggtgcct ga
//
