LOCUS       sequence-200310-        4391 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid encoding Sapphire.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4391)
  AUTHORS   Balleza E, Kim JM, Cluzel P
  TITLE     Systematic characterization of maturation time of fluorescent 
            proteins in living cells.
  JOURNAL   Nat Methods. 2018 Jan;15(1):47-51. doi: 10.1038/nmeth.4509. Epub 
            2017 Nov 20.
  PUBMED    29320486
REFERENCE   2  (bases 1 to 4391)
  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: 
            "10.1038/nmeth.4509"; journalName: "Nat Methods"; date: "2018-01"; 
            volume: "15"; issue: "1"; pages: "47-51"
COMMENT     Sequence Label: pEB1-Sapphire
FEATURES             Location/Qualifiers
     source          1..4391
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             194..910
                     /codon_start=1
                     /product="Aequorea victoria green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFSYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSIQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(224..252)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     847..867
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      948..1034
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(1572..2522)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      2570..2792
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(3284..3378)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(3409..4203)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3520..3539)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4130..4149
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     join(4382..4391,1..9)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcacgaggcc ctttcgtctt caccacagct aacaccacgt cgtccctatc tgctgcccta
       61 ggtctatgag tggttgctgg ataactttac gggcatgcat aaggctcgta tgatatattc
      121 agggagacca caacggtttc cctctacaaa taattttgtt taacttttct agatttaaga
      181 aggagatata catatgagta aaggagaaga acttttcact ggagttgtcc caattcttgt
      241 tgaattagat ggtgatgtta atgggcacaa attttctgtc agtggagagg gtgaaggtga
      301 tgcaacatac ggaaaactta cccttaaatt tatttgcact actggaaaac tacctgttcc
      361 atggccaaca cttgtcacta ctttctctta tggtgttcaa tgctttgcga gatacccaga
      421 tcatatgaaa cagcatgact ttttcaagag tgccatgccc gaaggttatg tacaggaaag
      481 aactatattt ttcaaagatg acgggaacta caagacacgt gctgaagtca agtttgaagg
      541 tgataccctt gttaatagaa tcgagttaaa aggtattgat tttaaagaag atggaaacat
      601 tcttggacac aaattggaat acaactttaa ctcacacaat gtatacatca tggcagacaa
      661 acaaaagaat ggaatcaaag ttaacttcaa aattagacac aacattgaag atggaagcgt
      721 tcaactagca gaccattatc aacaaaatac tccaattggc gatggccctg tccttttacc
      781 agacaaccat tacctgtcca tacaatctgc cctttcgaaa gatcccaacg aaaagagaga
      841 ccacatggtc cttcttgagt ttgtaacagc tgctgggatt acacatggca tggatgaact
      901 atacaaataa atgtccagac ctgcaggcat gcaagctcta gaggcatcaa ataaaacgaa
      961 aggctcagtc gaaagactgg gcctttcgtt ttatctgttg tttgtcggtg aacgctctcc
     1021 tgagtaggac aaatccgccg ccctagacct agggtacggg ttttgctgcc cgcaaacggg
     1081 ctgttctggt gttgctagtt tgttatcaga atcgcagatc cggcttcagc cggtttgccg
     1141 gctgaaagcg ctatttcttc cagaattgcc atgatttttt ccccacggga ggcgtcactg
     1201 gctcccgtgt tgtcggcagc tttgattcga taagcagcat cgcctgtttc aggctgtcta
     1261 tgtgtgactg ttgagctgta acaagttgtc tcaggtgttc aatttcatgt tctagttgct
     1321 ttgttttact ggtttcacct gttctattag gtgttacatg ctgttcatct gttacattgt
     1381 cgatctgttc atggtgaaca gctttgaatg caccaaaaac tcgtaaaagc tctgatgtat
     1441 ctatcttttt tacaccgttt tcatctgtgc atatggacag ttttcccttt gatatgtaac
     1501 ggtgaacagt tgttctactt ttgtttgtta gtcttgatgc ttcactgata gatacaagag
     1561 ccataagaac ctcagatcct tccgtattta gccagtatgt tctctagtgt ggttcgttgt
     1621 ttttgcgtga gccatgagaa cgaaccattg agatcatact tactttgcat gtcactcaaa
     1681 aattttgcct caaaactggt gagctgaatt tttgcagtta aagcatcgtg tagtgttttt
     1741 cttagtccgt tatgtaggta ggaatctgat gtaatggttg ttggtatttt gtcaccattc
     1801 atttttatct ggttgttctc aagttcggtt acgagatcca tttgtctatc tagttcaact
     1861 tggaaaatca acgtatcagt cgggcggcct cgcttatcaa ccaccaattt catattgctg
     1921 taagtgttta aatctttact tattggtttc aaaacccatt ggttaagcct tttaaactca
     1981 tggtagttat tttcaagcat taacatgaac ttaaattcat caaggctaat ctctatattt
     2041 gccttgtgag ttttcttttg tgttagttct tttaataacc actcataaat cctcatagag
     2101 tatttgtttt caaaagactt aacatgttcc agattatatt ttatgaattt ttttaactgg
     2161 aaaagataag gcaatatctc ttcactaaaa actaattcta atttttcgct tgagaacttg
     2221 gcatagtttg tccactggaa aatctcaaag cctttaacca aaggattcct gatttccaca
     2281 gttctcgtca tcagctctct ggttgcttta gctaatacac cataagcatt ttccctactg
     2341 atgttcatca tctgagcgta ttggttataa gtgaacgata ccgtccgttc tttccttgta
     2401 gggttttcaa tcgtggggtt gagtagtgcc acacagcata aaattagctt ggtttcatgc
     2461 tccgttaagt catagcgact aatcgctagt tcatttgctt tgaaaacaac taattcagac
     2521 atacatctca attggtctag gtgattttaa tcactatacc aattgagatg ggctagtcaa
     2581 tgataattac tagtcctttt cctttgagtt gtgggtatct gtaaattctg ctagaccttt
     2641 gctggaaaac ttgtaaattc tgctagaccc tctgtaaatt ccgctagacc tttgtgtgtt
     2701 ttttttgttt atattcaagt ggttataatt tatagaataa agaaagaata aaaaaagata
     2761 aaaagaatag atcccagccc tgtgtataac tcactacttt agtcagttcc gcagtattac
     2821 aaaaggatgt cgcaaacgct gtttgctcct ctacaaaaca gaccttaaaa ccctaaaggc
     2881 ttaagtagca ccctcgcaag ctcgggcaaa tcgctgaata ttccttttgt ctccgaccat
     2941 caggcacctg agtcgctgtc tttttcgtga cattcagttc gctgcgctca cggctctggc
     3001 agtgaatggg ggtaaatggc actacaggcg ccttttatgg attcatgcaa ggaaactacc
     3061 cataatacaa gaaaagcccg tcacgggctt ctcagggcgt tttatggcgg gtctgctatg
     3121 tggtgctatc tgactttttg ctgttcagca gttcctgccc tctgattttc cagtctgacc
     3181 acttcggatt atcccgtgac aggtcattca gactggctaa tgcacccagt aaggcagcgg
     3241 tatcatcaac aggcttaccc gtcttactgt ccctagtgct tggattctca ccaataaaaa
     3301 acgcccggcg gcaaccgagc gttctgaaca aatccagatg gagttctgag gtcattactg
     3361 gatctatcaa caggagtcca agcgagctct cgaaccccag agtcccgctc agaagaactc
     3421 gtcaagaagg cgatagaagg cgatgcgctg cgaatcggga gcggcgatac cgtaaagcac
     3481 gaggaagcgg tcagcccatt cgccgccaag ctcttcagca atatcacggg tagccaacgc
     3541 tatgtcctga tagcggtccg ccacacccag ccggccacag tcgatgaatc cagaaaagcg
     3601 gccattttcc accatgatat tcggcaagca ggcatcgcca tgggtcacga cgagatcctc
     3661 gccgtcgggc atgcgcgcct tgagcctggc gaacagttcg gctggcgcga gcccctgatg
     3721 ctcttcgtcc agatcatcct gatcgacaag accggcttcc atccgagtac gtgctcgctc
     3781 gatgcgatgt ttcgcttggt ggtcgaatgg gcaggtagcc ggatcaagcg tatgcagccg
     3841 ccgcattgca tcagccatga tggatacttt ctcggcagga gcaaggtgag atgacaggag
     3901 atcctgcccc ggcacttcgc ccaatagcag ccagtccctt cccgcttcag tgacaacgtc
     3961 gagcacagct gcgcaaggaa cgcccgtcgt ggccagccac gatagccgcg ctgcctcgtc
     4021 ctgcagttca ttcagggcac cggacaggtc ggtcttgaca aaaagaaccg ggcgcccctg
     4081 cgctgacagc cggaacacgg cggcatcaga gcagccgatt gtctgttgtg cccagtcata
     4141 gccgaatagc ctctccaccc aagcggccgg agaacctgcg tgcaatccat cttgttcaat
     4201 catgcgaaac gatcctcatc ctgtctcttg atcagatctt gatcccctgc gccatcagat
     4261 ccttggcggc aagaaagcca tccagtttac tttgcagggc ttcccaacct taccagaggg
     4321 cgccccagct ggcaattccg acgtctaaga aaccattatt atcatgacat taacctataa
     4381 aaataggcgt a
//
