LOCUS       sequence-200311-        4391 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid encoding T-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-T-Sapphire
FEATURES             Location/Qualifiers
     source          1..4391
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     58..76
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             261..977
                     /codon_start=1
                     /product="Aequorea victoria green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFSYGVMVFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYIMADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSIQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(291..319)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     914..934
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      1015..1101
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(1639..2589)
                     /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      2637..2859
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(3351..3445)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(3476..4270)
                     /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(3587..3606)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4197..4216
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 ccagctggca attccgacgt ctaagaaacc attattatca tgacattaac ctataaaaat
       61 aggcgtatca cgaggccctt tcgtcttcac cacagctaac accacgtcgt ccctatctgc
      121 tgccctaggt ctatgagtgg ttgctggata actttacggg catgcataag gctcgtatga
      181 tatattcagg gagaccacaa cggtttccct ctacaaataa ttttgtttaa cttttctaga
      241 tttaagaagg agatatacat atgagtaaag gagaagaact tttcactgga gttgtcccaa
      301 ttcttgttga attagatggt gatgttaatg ggcacaaatt ttctgtcagt ggagagggtg
      361 aaggtgatgc aacatacgga aaacttaccc ttaaatttat ttgcactact ggaaaactac
      421 ctgttccatg gccaacactt gtcactactt tctcttatgg tgttatggtc tttgcgagat
      481 acccagatca tatgaaacag catgactttt tcaagagtgc catgcccgaa ggttatgtac
      541 aggaaagaac tatatttttc aaagatgacg ggaactacaa gacacgtgct gaagtcaagt
      601 ttgaaggtga tacccttgtt aatagaatcg agttaaaagg tattgatttt aaagaagatg
      661 gaaacattct tggacacaaa ttggaataca actttaactc acacaatgta tacatcatgg
      721 cagacaaaca aaagaatgga atcaaagcga acttcaaaat tagacacaac attgaagatg
      781 gaggcgttca actagcagac cattatcaac aaaatactcc aattggcgat ggccctgtcc
      841 ttttaccaga caaccattac ctgtccatac aatctgccct ttcgaaagat cccaacgaaa
      901 agagagacca catggtcctt cttgagtttg taacagctgc tgggattaca catggcatgg
      961 atgaactata caaataaatg tccagacctg caggcatgca agctctagag gcatcaaata
     1021 aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac
     1081 gctctcctga gtaggacaaa tccgccgccc tagacctagg gtacgggttt tgctgcccgc
     1141 aaacgggctg ttctggtgtt gctagtttgt tatcagaatc gcagatccgg cttcagccgg
     1201 tttgccggct gaaagcgcta tttcttccag aattgccatg attttttccc cacgggaggc
     1261 gtcactggct cccgtgttgt cggcagcttt gattcgataa gcagcatcgc ctgtttcagg
     1321 ctgtctatgt gtgactgttg agctgtaaca agttgtctca ggtgttcaat ttcatgttct
     1381 agttgctttg ttttactggt ttcacctgtt ctattaggtg ttacatgctg ttcatctgtt
     1441 acattgtcga tctgttcatg gtgaacagct ttgaatgcac caaaaactcg taaaagctct
     1501 gatgtatcta tcttttttac accgttttca tctgtgcata tggacagttt tccctttgat
     1561 atgtaacggt gaacagttgt tctacttttg tttgttagtc ttgatgcttc actgatagat
     1621 acaagagcca taagaacctc agatccttcc gtatttagcc agtatgttct ctagtgtggt
     1681 tcgttgtttt tgcgtgagcc atgagaacga accattgaga tcatacttac tttgcatgtc
     1741 actcaaaaat tttgcctcaa aactggtgag ctgaattttt gcagttaaag catcgtgtag
     1801 tgtttttctt agtccgttat gtaggtagga atctgatgta atggttgttg gtattttgtc
     1861 accattcatt tttatctggt tgttctcaag ttcggttacg agatccattt gtctatctag
     1921 ttcaacttgg aaaatcaacg tatcagtcgg gcggcctcgc ttatcaacca ccaatttcat
     1981 attgctgtaa gtgtttaaat ctttacttat tggtttcaaa acccattggt taagcctttt
     2041 aaactcatgg tagttatttt caagcattaa catgaactta aattcatcaa ggctaatctc
     2101 tatatttgcc ttgtgagttt tcttttgtgt tagttctttt aataaccact cataaatcct
     2161 catagagtat ttgttttcaa aagacttaac atgttccaga ttatatttta tgaatttttt
     2221 taactggaaa agataaggca atatctcttc actaaaaact aattctaatt tttcgcttga
     2281 gaacttggca tagtttgtcc actggaaaat ctcaaagcct ttaaccaaag gattcctgat
     2341 ttccacagtt ctcgtcatca gctctctggt tgctttagct aatacaccat aagcattttc
     2401 cctactgatg ttcatcatct gagcgtattg gttataagtg aacgataccg tccgttcttt
     2461 ccttgtaggg ttttcaatcg tggggttgag tagtgccaca cagcataaaa ttagcttggt
     2521 ttcatgctcc gttaagtcat agcgactaat cgctagttca tttgctttga aaacaactaa
     2581 ttcagacata catctcaatt ggtctaggtg attttaatca ctataccaat tgagatgggc
     2641 tagtcaatga taattactag tccttttcct ttgagttgtg ggtatctgta aattctgcta
     2701 gacctttgct ggaaaacttg taaattctgc tagaccctct gtaaattccg ctagaccttt
     2761 gtgtgttttt tttgtttata ttcaagtggt tataatttat agaataaaga aagaataaaa
     2821 aaagataaaa agaatagatc ccagccctgt gtataactca ctactttagt cagttccgca
     2881 gtattacaaa aggatgtcgc aaacgctgtt tgctcctcta caaaacagac cttaaaaccc
     2941 taaaggctta agtagcaccc tcgcaagctc gggcaaatcg ctgaatattc cttttgtctc
     3001 cgaccatcag gcacctgagt cgctgtcttt ttcgtgacat tcagttcgct gcgctcacgg
     3061 ctctggcagt gaatgggggt aaatggcact acaggcgcct tttatggatt catgcaagga
     3121 aactacccat aatacaagaa aagcccgtca cgggcttctc agggcgtttt atggcgggtc
     3181 tgctatgtgg tgctatctga ctttttgctg ttcagcagtt cctgccctct gattttccag
     3241 tctgaccact tcggattatc ccgtgacagg tcattcagac tggctaatgc acccagtaag
     3301 gcagcggtat catcaacagg cttacccgtc ttactgtccc tagtgcttgg attctcacca
     3361 ataaaaaacg cccggcggca accgagcgtt ctgaacaaat ccagatggag ttctgaggtc
     3421 attactggat ctatcaacag gagtccaagc gagctctcga accccagagt cccgctcaga
     3481 agaactcgtc aagaaggcga tagaaggcga tgcgctgcga atcgggagcg gcgataccgt
     3541 aaagcacgag gaagcggtca gcccattcgc cgccaagctc ttcagcaata tcacgggtag
     3601 ccaacgctat gtcctgatag cggtccgcca cacccagccg gccacagtcg atgaatccag
     3661 aaaagcggcc attttccacc atgatattcg gcaagcaggc atcgccatgg gtcacgacga
     3721 gatcctcgcc gtcgggcatg cgcgccttga gcctggcgaa cagttcggct ggcgcgagcc
     3781 cctgatgctc ttcgtccaga tcatcctgat cgacaagacc ggcttccatc cgagtacgtg
     3841 ctcgctcgat gcgatgtttc gcttggtggt cgaatgggca ggtagccgga tcaagcgtat
     3901 gcagccgccg cattgcatca gccatgatgg atactttctc ggcaggagca aggtgagatg
     3961 acaggagatc ctgccccggc acttcgccca atagcagcca gtcccttccc gcttcagtga
     4021 caacgtcgag cacagctgcg caaggaacgc ccgtcgtggc cagccacgat agccgcgctg
     4081 cctcgtcctg cagttcattc agggcaccgg acaggtcggt cttgacaaaa agaaccgggc
     4141 gcccctgcgc tgacagccgg aacacggcgg catcagagca gccgattgtc tgttgtgccc
     4201 agtcatagcc gaatagcctc tccacccaag cggccggaga acctgcgtgc aatccatctt
     4261 gttcaatcat gcgaaacgat cctcatcctg tctcttgatc agatcttgat cccctgcgcc
     4321 atcagatcct tggcggcaag aaagccatcc agtttacttt gcagggcttc ccaaccttac
     4381 cagagggcgc c
//
