LOCUS       sequence-200315-        4391 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid encoding mGFPmut2.
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-mGFPmut2
FEATURES             Location/Qualifiers
     source          1..4391
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     9..27
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             212..925
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFAYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(242..270)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     865..885
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      966..1052
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(1590..2540)
                     /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      2588..2810
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(3302..3396)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(3427..4221)
                     /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(3538..3557)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4148..4167
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 cctataaaaa taggcgtatc acgaggccct ttcgtcttca ccacagctaa caccacgtcg
       61 tccctatctg ctgccctagg tctatgagtg gttgctggat aactttacgg gcatgcataa
      121 ggctcgtatg atatattcag ggagaccaca acggtttccc tctacaaata attttgttta
      181 acttttctag atttaagaag gagatataca tatgagtaaa ggagaagaac ttttcactgg
      241 agttgtccca attcttgttg aattagatgg tgatgttaat gggcacaaat tttctgtcag
      301 tggagagggt gaaggtgatg caacatacgg aaaacttacc cttaaattta tttgcactac
      361 tggaaaacta cctgttccat ggccaacact tgtcactact ttcgcgtatg gtcttcaatg
      421 ctttgcgaga tacccagatc atatgaaaca gcatgacttt ttcaagagtg ccatgcccga
      481 aggttatgta caggaaagaa ctatattttt caaagatgac gggaactaca agacacgtgc
      541 tgaagtcaag tttgaaggtg atacccttgt taatagaatc gagttaaaag gtattgattt
      601 taaagaagat ggaaacattc ttggacacaa attggaatac aactataact cacacaatgt
      661 atacatcatg gcagacaaac aaaagaatgg aatcaaagtt aacttcaaaa ttagacacaa
      721 cattgaagat ggaagcgttc aactagcaga ccattatcaa caaaatactc caattggcga
      781 tggccctgtc cttttaccag acaaccatta cctgtccaca caatctaagc tttcgaaaga
      841 tcccaacgaa aagagagacc acatggtcct tcttgagttt gtaacagctg ctgggattac
      901 acatggcatg gatgaactat acaaataaat gtccagacct gcaggcatgc aagctctaga
      961 ggcatcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt
     1021 tgtcggtgaa cgctctcctg agtaggacaa atccgccgcc ctagacctag ggtacgggtt
     1081 ttgctgcccg caaacgggct gttctggtgt tgctagtttg ttatcagaat cgcagatccg
     1141 gcttcagccg gtttgccggc tgaaagcgct atttcttcca gaattgccat gattttttcc
     1201 ccacgggagg cgtcactggc tcccgtgttg tcggcagctt tgattcgata agcagcatcg
     1261 cctgtttcag gctgtctatg tgtgactgtt gagctgtaac aagttgtctc aggtgttcaa
     1321 tttcatgttc tagttgcttt gttttactgg tttcacctgt tctattaggt gttacatgct
     1381 gttcatctgt tacattgtcg atctgttcat ggtgaacagc tttgaatgca ccaaaaactc
     1441 gtaaaagctc tgatgtatct atctttttta caccgttttc atctgtgcat atggacagtt
     1501 ttccctttga tatgtaacgg tgaacagttg ttctactttt gtttgttagt cttgatgctt
     1561 cactgataga tacaagagcc ataagaacct cagatccttc cgtatttagc cagtatgttc
     1621 tctagtgtgg ttcgttgttt ttgcgtgagc catgagaacg aaccattgag atcatactta
     1681 ctttgcatgt cactcaaaaa ttttgcctca aaactggtga gctgaatttt tgcagttaaa
     1741 gcatcgtgta gtgtttttct tagtccgtta tgtaggtagg aatctgatgt aatggttgtt
     1801 ggtattttgt caccattcat ttttatctgg ttgttctcaa gttcggttac gagatccatt
     1861 tgtctatcta gttcaacttg gaaaatcaac gtatcagtcg ggcggcctcg cttatcaacc
     1921 accaatttca tattgctgta agtgtttaaa tctttactta ttggtttcaa aacccattgg
     1981 ttaagccttt taaactcatg gtagttattt tcaagcatta acatgaactt aaattcatca
     2041 aggctaatct ctatatttgc cttgtgagtt ttcttttgtg ttagttcttt taataaccac
     2101 tcataaatcc tcatagagta tttgttttca aaagacttaa catgttccag attatatttt
     2161 atgaattttt ttaactggaa aagataaggc aatatctctt cactaaaaac taattctaat
     2221 ttttcgcttg agaacttggc atagtttgtc cactggaaaa tctcaaagcc tttaaccaaa
     2281 ggattcctga tttccacagt tctcgtcatc agctctctgg ttgctttagc taatacacca
     2341 taagcatttt ccctactgat gttcatcatc tgagcgtatt ggttataagt gaacgatacc
     2401 gtccgttctt tccttgtagg gttttcaatc gtggggttga gtagtgccac acagcataaa
     2461 attagcttgg tttcatgctc cgttaagtca tagcgactaa tcgctagttc atttgctttg
     2521 aaaacaacta attcagacat acatctcaat tggtctaggt gattttaatc actataccaa
     2581 ttgagatggg ctagtcaatg ataattacta gtccttttcc tttgagttgt gggtatctgt
     2641 aaattctgct agacctttgc tggaaaactt gtaaattctg ctagaccctc tgtaaattcc
     2701 gctagacctt tgtgtgtttt ttttgtttat attcaagtgg ttataattta tagaataaag
     2761 aaagaataaa aaaagataaa aagaatagat cccagccctg tgtataactc actactttag
     2821 tcagttccgc agtattacaa aaggatgtcg caaacgctgt ttgctcctct acaaaacaga
     2881 ccttaaaacc ctaaaggctt aagtagcacc ctcgcaagct cgggcaaatc gctgaatatt
     2941 ccttttgtct ccgaccatca ggcacctgag tcgctgtctt tttcgtgaca ttcagttcgc
     3001 tgcgctcacg gctctggcag tgaatggggg taaatggcac tacaggcgcc ttttatggat
     3061 tcatgcaagg aaactaccca taatacaaga aaagcccgtc acgggcttct cagggcgttt
     3121 tatggcgggt ctgctatgtg gtgctatctg actttttgct gttcagcagt tcctgccctc
     3181 tgattttcca gtctgaccac ttcggattat cccgtgacag gtcattcaga ctggctaatg
     3241 cacccagtaa ggcagcggta tcatcaacag gcttacccgt cttactgtcc ctagtgcttg
     3301 gattctcacc aataaaaaac gcccggcggc aaccgagcgt tctgaacaaa tccagatgga
     3361 gttctgaggt cattactgga tctatcaaca ggagtccaag cgagctctcg aaccccagag
     3421 tcccgctcag aagaactcgt caagaaggcg atagaaggcg atgcgctgcg aatcgggagc
     3481 ggcgataccg taaagcacga ggaagcggtc agcccattcg ccgccaagct cttcagcaat
     3541 atcacgggta gccaacgcta tgtcctgata gcggtccgcc acacccagcc ggccacagtc
     3601 gatgaatcca gaaaagcggc cattttccac catgatattc ggcaagcagg catcgccatg
     3661 ggtcacgacg agatcctcgc cgtcgggcat gcgcgccttg agcctggcga acagttcggc
     3721 tggcgcgagc ccctgatgct cttcgtccag atcatcctga tcgacaagac cggcttccat
     3781 ccgagtacgt gctcgctcga tgcgatgttt cgcttggtgg tcgaatgggc aggtagccgg
     3841 atcaagcgta tgcagccgcc gcattgcatc agccatgatg gatactttct cggcaggagc
     3901 aaggtgagat gacaggagat cctgccccgg cacttcgccc aatagcagcc agtcccttcc
     3961 cgcttcagtg acaacgtcga gcacagctgc gcaaggaacg cccgtcgtgg ccagccacga
     4021 tagccgcgct gcctcgtcct gcagttcatt cagggcaccg gacaggtcgg tcttgacaaa
     4081 aagaaccggg cgcccctgcg ctgacagccg gaacacggcg gcatcagagc agccgattgt
     4141 ctgttgtgcc cagtcatagc cgaatagcct ctccacccaa gcggccggag aacctgcgtg
     4201 caatccatct tgttcaatca tgcgaaacga tcctcatcct gtctcttgat cagatcttga
     4261 tcccctgcgc catcagatcc ttggcggcaa gaaagccatc cagtttactt tgcagggctt
     4321 cccaacctta ccagagggcg ccccagctgg caattccgac gtctaagaaa ccattattat
     4381 catgacatta a
//
