LOCUS       sequence-200469-        4391 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid encoding mClover3.
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-mClover3
FEATURES             Location/Qualifiers
     source          1..4391
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     185..204
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     437..455
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             640..1353
                     /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="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHYVYITADKQKNCIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(670..698)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1293..1313
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      1394..1480
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2018..2968)
                     /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      3016..3238
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(3730..3824)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(join(3855..4391,1..258))
                     /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(3966..3985)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
ORIGIN
        1 ttcagtgaca acgtcgagca cagctgcgca aggaacgccc gtcgtggcca gccacgatag
       61 ccgcgctgcc tcgtcctgca gttcattcag ggcaccggac aggtcggtct tgacaaaaag
      121 aaccgggcgc ccctgcgctg acagccggaa cacggcggca tcagagcagc cgattgtctg
      181 ttgtgcccag tcatagccga atagcctctc cacccaagcg gccggagaac ctgcgtgcaa
      241 tccatcttgt tcaatcatgc gaaacgatcc tcatcctgtc tcttgatcag atcttgatcc
      301 cctgcgccat cagatccttg gcggcaagaa agccatccag tttactttgc agggcttccc
      361 aaccttacca gagggcgccc cagctggcaa ttccgacgtc taagaaacca ttattatcat
      421 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtcttcacc acagctaaca
      481 ccacgtcgtc cctatctgct gccctaggtc tatgagtggt tgctggataa ctttacgggc
      541 atgcataagg ctcgtatgat atattcaggg agaccacaac ggtttccctc tacaaataat
      601 tttgtttaac ttttctagat ttaagaagga gatatacata tgagtaaagg agaagaactt
      661 ttcactggag ttgtcccaat tcttgttgaa ttagatggtg atgttaatgg gcacaaattt
      721 tctgtcagag gagagggtga aggtgatgca acaaacggaa aacttaccct taaatttatt
      781 tgcactactg gaaaactacc tgttccatgg ccaacacttg tcactacttt cggttatggt
      841 gttgcatgct tttcaagata cccagatcat atgaaacagc atgacttttt caagagtgcc
      901 atgcccgaag gttatgtaca ggaaagaact atatctttca aagatgacgg gacctacaag
      961 acacgtgctg aagtcaagtt tgaaggtgat acccttgtta atagaatcga gttaaaaggt
     1021 attgatttta aagaagatgg aaacattctt ggacacaaat tggaatacaa ctttaactca
     1081 cactatgtat acatcacggc agacaaacaa aagaattgca tcaaagctaa cttcaaaatt
     1141 agacacaacg ttgaagatgg aagcgttcaa ctagcagacc attatcaaca aaatactcca
     1201 attggcgatg gccctgtcct tttaccagac aaccattacc tgtcccacca atctaagctt
     1261 tcgaaagatc ccaacgaaaa gagagaccac atggtccttc ttgagtttgt aacagctgct
     1321 gggattacac atggcatgga tgaactatac aaataaatgt ccagacctgc aggcatgcaa
     1381 gctctagagg catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat
     1441 ctgttgtttg tcggtgaacg ctctcctgag taggacaaat ccgccgccct agacctaggg
     1501 tacgggtttt gctgcccgca aacgggctgt tctggtgttg ctagtttgtt atcagaatcg
     1561 cagatccggc ttcagccggt ttgccggctg aaagcgctat ttcttccaga attgccatga
     1621 ttttttcccc acgggaggcg tcactggctc ccgtgttgtc ggcagctttg attcgataag
     1681 cagcatcgcc tgtttcaggc tgtctatgtg tgactgttga gctgtaacaa gttgtctcag
     1741 gtgttcaatt tcatgttcta gttgctttgt tttactggtt tcacctgttc tattaggtgt
     1801 tacatgctgt tcatctgtta cattgtcgat ctgttcatgg tgaacagctt tgaatgcacc
     1861 aaaaactcgt aaaagctctg atgtatctat cttttttaca ccgttttcat ctgtgcatat
     1921 ggacagtttt ccctttgata tgtaacggtg aacagttgtt ctacttttgt ttgttagtct
     1981 tgatgcttca ctgatagata caagagccat aagaacctca gatccttccg tatttagcca
     2041 gtatgttctc tagtgtggtt cgttgttttt gcgtgagcca tgagaacgaa ccattgagat
     2101 catacttact ttgcatgtca ctcaaaaatt ttgcctcaaa actggtgagc tgaatttttg
     2161 cagttaaagc atcgtgtagt gtttttctta gtccgttatg taggtaggaa tctgatgtaa
     2221 tggttgttgg tattttgtca ccattcattt ttatctggtt gttctcaagt tcggttacga
     2281 gatccatttg tctatctagt tcaacttgga aaatcaacgt atcagtcggg cggcctcgct
     2341 tatcaaccac caatttcata ttgctgtaag tgtttaaatc tttacttatt ggtttcaaaa
     2401 cccattggtt aagcctttta aactcatggt agttattttc aagcattaac atgaacttaa
     2461 attcatcaag gctaatctct atatttgcct tgtgagtttt cttttgtgtt agttctttta
     2521 ataaccactc ataaatcctc atagagtatt tgttttcaaa agacttaaca tgttccagat
     2581 tatattttat gaattttttt aactggaaaa gataaggcaa tatctcttca ctaaaaacta
     2641 attctaattt ttcgcttgag aacttggcat agtttgtcca ctggaaaatc tcaaagcctt
     2701 taaccaaagg attcctgatt tccacagttc tcgtcatcag ctctctggtt gctttagcta
     2761 atacaccata agcattttcc ctactgatgt tcatcatctg agcgtattgg ttataagtga
     2821 acgataccgt ccgttctttc cttgtagggt tttcaatcgt ggggttgagt agtgccacac
     2881 agcataaaat tagcttggtt tcatgctccg ttaagtcata gcgactaatc gctagttcat
     2941 ttgctttgaa aacaactaat tcagacatac atctcaattg gtctaggtga ttttaatcac
     3001 tataccaatt gagatgggct agtcaatgat aattactagt ccttttcctt tgagttgtgg
     3061 gtatctgtaa attctgctag acctttgctg gaaaacttgt aaattctgct agaccctctg
     3121 taaattccgc tagacctttg tgtgtttttt ttgtttatat tcaagtggtt ataatttata
     3181 gaataaagaa agaataaaaa aagataaaaa gaatagatcc cagccctgtg tataactcac
     3241 tactttagtc agttccgcag tattacaaaa ggatgtcgca aacgctgttt gctcctctac
     3301 aaaacagacc ttaaaaccct aaaggcttaa gtagcaccct cgcaagctcg ggcaaatcgc
     3361 tgaatattcc ttttgtctcc gaccatcagg cacctgagtc gctgtctttt tcgtgacatt
     3421 cagttcgctg cgctcacggc tctggcagtg aatgggggta aatggcacta caggcgcctt
     3481 ttatggattc atgcaaggaa actacccata atacaagaaa agcccgtcac gggcttctca
     3541 gggcgtttta tggcgggtct gctatgtggt gctatctgac tttttgctgt tcagcagttc
     3601 ctgccctctg attttccagt ctgaccactt cggattatcc cgtgacaggt cattcagact
     3661 ggctaatgca cccagtaagg cagcggtatc atcaacaggc ttacccgtct tactgtccct
     3721 agtgcttgga ttctcaccaa taaaaaacgc ccggcggcaa ccgagcgttc tgaacaaatc
     3781 cagatggagt tctgaggtca ttactggatc tatcaacagg agtccaagcg agctctcgaa
     3841 ccccagagtc ccgctcagaa gaactcgtca agaaggcgat agaaggcgat gcgctgcgaa
     3901 tcgggagcgg cgataccgta aagcacgagg aagcggtcag cccattcgcc gccaagctct
     3961 tcagcaatat cacgggtagc caacgctatg tcctgatagc ggtccgccac acccagccgg
     4021 ccacagtcga tgaatccaga aaagcggcca ttttccacca tgatattcgg caagcaggca
     4081 tcgccatggg tcacgacgag atcctcgccg tcgggcatgc gcgccttgag cctggcgaac
     4141 agttcggctg gcgcgagccc ctgatgctct tcgtccagat catcctgatc gacaagaccg
     4201 gcttccatcc gagtacgtgc tcgctcgatg cgatgtttcg cttggtggtc gaatgggcag
     4261 gtagccggat caagcgtatg cagccgccgc attgcatcag ccatgatgga tactttctcg
     4321 gcaggagcaa ggtgagatga caggagatcc tgccccggca cttcgcccaa tagcagccag
     4381 tcccttcccg c
//
