LOCUS       sequence-481220-        6990 bp    DNA     circular SYN 30-JUL-2025
DEFINITION  TurboRFP constitutive Golden Gate construct.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6990)
  TITLE     Murray Lab Golden Gate Plasmids 2025
REFERENCE   2  (bases 1 to 6990)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 30, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: eOYW002
FEATURES             Location/Qualifiers
     source          1..6990
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(171..962,963..1031))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     794..813
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1032..1136)
                     /gene="bla"
                     /label=AmpR promoter
     mobile_element  1210..1375
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     primer_bind     complement(1494..1513)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1505..2320
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             1505..2320
                     /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="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1578..1597)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4034..4052
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4119..4153
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     RBS             4162..4173
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             4180..4875
                     /codon_start=1
                     /product="red fluorescent protein from Entacmaea 
                     quadricolor"
                     /label=TurboRFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMKIKV
                     VEGGPLPFAFDILATSFMYGSKAFINHTQGIPDFFKQSFPEGFTWERITTYEDGGVLTA
                     TQDTSFQNGCIIYNVKINGVNFPSNGPVMQKKTRGWEANTEMLYPADGGLRGHSQMALK
                     LVGGGYLHCSFKTTYRSKKPAKNLKMPGFHFVDHRLERIKEADKETYVEQHEMAVAKYC
                     DLPSKLGHR"
     terminator      4880..4974
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      5379..5767
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     CDS             complement(5938..6309)
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     oriT            6342..6451
                     /note="incP origin of transfer"
ORIGIN
        1 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
       61 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
      121 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
      181 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
      241 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
      301 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
      361 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
      421 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
      481 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
      541 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
      601 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
      661 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
      721 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
      781 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
      841 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
      901 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
      961 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     1021 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     1081 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     1141 tttccccgaa aagtgccacc tgaattaatt cgagctcgcc caaacataac aggaagaaaa
     1201 atgccccgct gtgggcggac aaaatagttg ggaactggga ggggtggaaa tggagttttt
     1261 aaggattatt tagggaagag tgacaaaata gatgggaact gggtgtagcg tcgtaagcta
     1321 atacgaaaat taaaaatgac aaaatagttt ggaactagat ttcacttatc tggttggcct
     1381 gcaggggggg ggggaaagcc acgttgtgtc tcaaaatctc tgatgttaca ttgcacaaga
     1441 taaaaatata tcatcatgaa caataaaact gtctgcttac ataaacagta atacaagggg
     1501 tgttatgagc catattcaac gggaaacgtc ttgctcgagg ccgcgattaa attccaacat
     1561 ggatgctgat ttatatgggt ataaatgggc tcgcgataat gtcgggcaat caggtgcgac
     1621 aatctatcga ttgtatggga agcccgatgc gccagagttg tttctgaaac atggcaaagg
     1681 tagcgttgcc aatgatgtta cagatgagat ggtcagacta aactggctga cggaatttat
     1741 gcctcttccg accatcaagc attttatccg tactcctgat gatgcatggt tactcaccac
     1801 tgcgatcccc gggaaaacag cattccaggt attagaagaa tatcctgatt caggtgaaaa
     1861 tattgttgat gcgctggcag tgttcctgcg ccggttgcat tcgattcctg tttgtaattg
     1921 tccttttaac agcgatcgcg tatttcgtct cgctcaggcg caatcacgaa tgaataacgg
     1981 tttggttgat gcgagtgatt ttgatgacga gcgtaatggc tggcctgttg aacaagtctg
     2041 gaaagaaatg cataagcttt tgccattctc accggattca gtcgtcactc atggtgattt
     2101 ctcacttgat aaccttattt ttgacgaggg gaaattaata ggttgtattg atgttggacg
     2161 agtcggaatc gcagaccgat accaggatct tgccatccta tggaactgcc tcggtgagtt
     2221 ttctccttca ttacagaaac ggctttttca aaaatatggt attgataatc ctgatatgaa
     2281 taaattgcag tttcatttga tgctcgatga gtttttctaa tcagaattgg ttaattggtt
     2341 gtaacactgg cagagcatta cgctgacttg acgggacggc ggctttgttg aataaatcga
     2401 acttttgctg agttgaagga tcagatcacg catcttcccg acaacgcaga ccgttccgtg
     2461 gcaaagcaaa agttcaaaat caccaactgg tccacctaca acaaagctct catcaaccgt
     2521 ggctccctca ctttctggct ggatgatggg gcgattcagg cctggtatga gtcagcaaca
     2581 ccttcttcac gaggcagacc tcagcgcccc cccccccctg caggtcgact agagtcgggg
     2641 atccggtgat tgattgagca agctttatgc ttgtaaaccg ttttgtgaaa aaatttttaa
     2701 aataaaaaag gggacctcta gggtccccaa ttaattagta atataatcta ttaaaggtca
     2761 ttcaaaaggt catccaccgg atcaattccc ctgctcgcgc aggctgggtg ccaagctctc
     2821 gggtaacatc aaggcccgat ccttggagcc cttgccctcc cgcacgatga tcgtgccgtg
     2881 atcgaaatcc agatccttga cccgcagttg caaaccctca ctgatccgca tgcccgttcc
     2941 atacagaagc tgggcgaaca aacgatgctc gccttccaga aaaccgagga tgcgaaccac
     3001 ttcatccggg gtcagcacca ccggcaagcg ccgcgacggc cgaggtcttc cgatctcctg
     3061 aagccagggc agatccgtgc acagcacctt gccgtagaag aacagcaagg ccgccaatgc
     3121 ctgacgatgc gtggagaccg aaaccttgcg ctcgttcgcc agccaggaca gaaatgcctc
     3181 gacttcgctg ctgcccaagg ttgccgggtg acgcacaccg tggaaacgga tgaaggcacg
     3241 aacccagtgg acataagcct gttcggttcg taagctgtaa tgcaagtagc gtatgcgctc
     3301 acgcaactgg tccagaacct tgaccgaacg cagcggtggt aacggcgcag tggcggtttt
     3361 catggcttgt tatgactgtt tttttggggt acagtctatg cctcgggcat ccaagcagca
     3421 agcgcgttac gccgtgggtc gatgtttgtg tctaacaatt cgttcaagcc gacgccgctt
     3481 cgcggcgcgg cttaactcaa gcgttagatg cactaagcac ataattgctc acagccaaac
     3541 tatcaggtca agtctgcttt tattattttt aagcgtgcat aataagccct acacaaattg
     3601 ggagatatat catgaaaggc tggctttttc ttgttatcgc aatagttggc gaagtaatcg
     3661 caacatccgc attaaaatct agcgagggct ttactaagct gatccggtgg atgacctttt
     3721 gaatgacctt taatagatta tattactaat taattgggga ccctagaggt cccctttttt
     3781 attttaaaaa ttttttcaca aaacggttta caagcataaa gcttgctcaa tcaatcaccg
     3841 gatcccgacg ggcccggtac cgcggccgcg cgaattcgag ctcggtacca tttatcaggg
     3901 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     3961 tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac
     4021 attaacctat aaaaataggc gtatcacgag gccctttcgt cttcacctcg agaacattac
     4081 tcgcatccat tctcaggctg tctcgtctcg tctcggagtt tacagctagc tcagtcctag
     4141 gtattatgct agctactaga gaaagaggag aaatactaaa tgagcgagct gatcaaggag
     4201 aacatgcaca tgaagctgta catggagggc accgtgaaca accaccactt caagtgcaca
     4261 tccgagggcg aaggcaagcc ctacgagggc acccagacca tgaagatcaa ggtggtcgag
     4321 ggcggccctc tccccttcgc cttcgacatc ctggctacca gcttcatgta cggcagcaaa
     4381 gccttcatca accacaccca gggcatcccc gacttcttta agcagtcctt ccctgagggc
     4441 ttcacatggg agagaatcac cacatacgaa gacgggggcg tgctgaccgc tacccaggac
     4501 accagcttcc agaacggctg catcatctac aacgtcaaga tcaacggggt gaacttccca
     4561 tccaacggcc ctgtgatgca gaagaaaaca cgcggctggg aggccaacac cgagatgctg
     4621 taccccgctg acggcggcct gagaggccac agccagatgg ccctgaagct cgtgggcggg
     4681 ggctacctgc actgctcctt caagaccaca tacagatcca agaaacccgc taagaacctc
     4741 aagatgcccg gcttccactt cgtggaccac agactggaaa gaatcaagga ggccgacaaa
     4801 gaaacgtacg tcgagcagca cgagatggct gtggccaagt actgcgacct ccctagcaaa
     4861 ctggggcaca gatgaaggtg actcctgttg atagatccag taatgacctc agaactccat
     4921 ctggatttgt tcagaacgct cggttgccgc cgggcgtttt ttattggtga gaatgcttcc
     4981 aggatacata gattaccaca actccgagcc cttccacccc aagctagctt ggaggacaaa
     5041 tccgccgcta ggagcttgcg gccgccttaa gacgcgtgga tccccgggaa tttcgacccc
     5101 acgcccctct ttaatacgac gggcaatttg cacttcagaa aatgaagagt ttgctttagc
     5161 cataacaaaa gtccagtatg ctttttcaca gcataactgg actgatttca gtttacaact
     5221 attctgtcta gtttaagact ttattgtcat agtttagatc tattttgttc agtttaagac
     5281 tttattgtcc gcccacaccc gcttacgcag ggcatccatt tattactcaa ccgtaaccga
     5341 ttttgccagg ttacgcggct ggtcctctag aattcccatg tcagccgtta agtgttcctg
     5401 tgtcactcaa aattgctttg agaggctcta agggcttctc agtgcgttac atccctggct
     5461 tgttgtccac aaccgttaaa ccttaaaagc tttaaaagcc ttatatattc ttttttttct
     5521 tataaaactt aaaaccttag aggctattta agttgctgat ttatattaat tttattgttc
     5581 aaacatgaga gcttagtacg tgaaacatga gagcttagta cgttagccat gagagcttag
     5641 tacgttagcc atgagggttt agttcgttaa acatgagagc ttagtacgtt aaacatgaga
     5701 gcttagtacg tgaaacatga gagcttagta cgtactatca acaggttgaa ctgctgatct
     5761 tcagatcctc tacgccggac gcatcgtggc cggatcgggg ggggggggcg ctcggtcttg
     5821 ccttgctcgt cggtgatgta cttcaccagc tccgcgaagt cgctcttctt gatggagcgc
     5881 atggggacgt gcttggcaat cacgcgcacc ccccggccgt tttagcggct aaaaaagtca
     5941 tggctctgcc ctcgggcgga ccacgcccat catgaccttg ccaagctcgt cctgcttctc
     6001 ttcgatcttc gccagcaggg cgaggatcgt ggcatcaccg aaccgcgccg tgcgcgggtc
     6061 gtcggtgagc cagagtttca gcaggccgcc caggcggccc aggtcgccat tgatgcgggc
     6121 cagctcgcgg acgtgctcat agtccacgac gcccgtgatt ttgtagccct ggccgacggc
     6181 cagcaggtag gccgacaggc tcatgccggc cgccgccgcc ttttcctcaa tcgctcttcg
     6241 ttcgtctgga aggcagtaca ccttgatagg tgggctgccc ttcctggttg gcttggtttc
     6301 atcagccatc cgcttgccct catctgttac gccggcggta gccggccagc ctcgcagagc
     6361 aggattcccg ttgagcaccg ccaggtgcga ataagggaca gtgaagaagg aacacccgct
     6421 cgcgggtggg cctacttcac ctatcctgcc cggctgacgc cgttggatac accaaggaaa
     6481 gtctacacga accctttggc aaaatcctgt atatcgtgcg aaaaaggatg gatataccga
     6541 aaaaatcgct ataatgaccc cgaagcaggg ttatgcagcg gaaaagcgct ttctcatagc
     6601 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     6661 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     6721 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     6781 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     6841 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     6901 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     6961 cagattacgc gcagaaaaaa aggatctcaa
//
