LOCUS       sequence-147997-        4731 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Localization: C1 Cloning Vector, Excitation: 399, Emission: 511.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4731)
  AUTHORS   Ai HW, Hazelwood KL, Davidson MW, Campbell RE
  TITLE     Fluorescent protein FRET pairs for ratiometric imaging of dual 
            biosensors.
  JOURNAL   Nat Methods. 2008 May;5(5):401-3. doi: 10.1038/nmeth.1207.
  PUBMED    18425137
REFERENCE   2  (bases 1 to 4731)
  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"; 
            journalName: "Nat Methods"; date: "2008-05"; volume: "5"; issue: 
            "5"; pages: "401-3"
COMMENT     Sequence Label: mT-Sapphire-C1
FEATURES             Location/Qualifiers
     source          1..4731
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        68..371
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        372..575
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     526..546
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      614..623
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(620..622,623..625,626..1336)
                     /codon_start=1
                     /product="strongly enhanced green fluorescent protein 
                     (Kremers et al., 2007)"
                     /label=SGFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFSYGVMVFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSIQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(665..686)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(926..945)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1273..1294
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    1337..1402
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    1526..1647
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1563..1582)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1617..1636
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1654..2109)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1791..1812)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2003..2022
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2136..2240
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2242..2599
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2242..2433
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2265..2285)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2450..2585
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2634..3428
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2634..3428
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2688..2707)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3298..3317
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3616..3635)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3660..3707
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4036..4624
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4525..4544
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 catgcattag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg
       61 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc
      121 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt
      181 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc
      241 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg
      301 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg
      361 ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact
      421 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa
      481 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta
      541 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct
      601 agcgctaccg gtcgccacca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
      661 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
      721 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct
      781 gcccgtgccc tggcccaccc tcgtgaccac cttctcctac ggcgtgatgg tgttcgcccg
      841 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
      901 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
      961 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
     1021 cggcaacatc ctggggcaca agctggagta caacttcaac agccacaacg tctatatcat
     1081 ggccgacaag cagaagaacg gcatcaaggc caacttcaag atccgccaca acatcgagga
     1141 cggcggcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
     1201 gctgctgccc gacaaccact acctgagcat ccagtccaag ctgagcaaag accccaacga
     1261 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctgggcat
     1321 ggacgagctg tacaagtccg gactcagatc tcgagctcaa gcttcgaatt ctgcagtcga
     1381 cggtaccgcg ggcccgggat ccaccggatc tagataactg atcataatca gccataccac
     1441 atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca
     1501 taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata
     1561 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     1621 tttgtccaaa ctcatcaatg tatcttaacg cgtaaattgt aagcgttaat attttgttaa
     1681 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     1741 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     1801 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     1861 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
     1921 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     1981 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
     2041 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     2101 agggcgcgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     2161 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     2221 aatattgaaa aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt
     2281 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     2341 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     2401 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     2461 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     2521 agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg
     2581 aggcctaggc ttttgcaaag atcgatcaag agacaggatg aggatcgttt cgcatgattg
     2641 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg
     2701 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg
     2761 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg
     2821 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg
     2881 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc
     2941 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc
     3001 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc
     3061 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc
     3121 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg
     3181 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct
     3241 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt
     3301 tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc
     3361 tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt
     3421 tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc
     3481 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     3541 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     3601 tagggggagg ctaactgaaa cacggaagga gacaataccg gaaggaaccc gcgctatgac
     3661 ggcaataaaa agacagaata aaacgcacgg tgttgggtcg tttgttcata aacgcggggt
     3721 tcggtcccag ggctggcact ctgtcgatac cccaccgaga ccccattggg gccaatacgc
     3781 ccgcgtttct tccttttccc caccccaccc cccaagttcg ggtgaaggcc cagggctcgc
     3841 agccaacgtc ggggcggcag gccctgccat agcctcaggt tactcatata tactttagat
     3901 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     3961 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     4021 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     4081 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     4141 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     4201 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     4261 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     4321 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     4381 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     4441 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     4501 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     4561 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     4621 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     4681 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg c
//
