LOCUS       sequence-100637-        6042 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  GFP-labeled SAH scaffold with split GFP attachment site.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6042)
  AUTHORS   Norris SR, Soppina V, Dizaji AS, Schimert KI, Sept D, Cai D, 
            Sivaramakrishnan S, Verhey KJ
  TITLE     A method for multiprotein assembly in cells reveals independent 
            action of kinesins in complex.
  JOURNAL   J Cell Biol. 2014 Nov 10;207(3):393-406. doi: 10.1083/jcb.201407086.
            Epub 2014 Nov 3.
  PUBMED    25365993
REFERENCE   2  (bases 1 to 6042)
  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: 
            "10.1083/jcb.201407086"; journalName: "J Cell Biol"; date: 
            "2014-11-10- 10"; volume: "207"; issue: "3"; pages: "393-406"
COMMENT     Sequence Label: mEGFP-30nmSAH-GFP(1-10)
FEATURES             Location/Qualifiers
     source          1..6042
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(658..679)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(919..938)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1266..1287
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2032..2673
                     /codon_start=1
                     /product="fragment of superfolder GFP consisting of the 
                     first 10 beta-strands"
                     /label=GFP(1-10)
                     /note="associates with GFP11, the 11th beta-strand, to 
                     reconstitute fluorescent GFP (Cabantous et al., 2005; 
                     Kamiyama et al., 2016)"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATIGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     KYKTRAVVKFEGDTLVNRIELKGTDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFTVRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQTVLSKDPNEK"
     polyA_signal    2830..2951
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2867..2886)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2921..2940
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2958..3413)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3095..3116)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3307..3326
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3440..3544
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3546..3903
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3546..3737
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3569..3589)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3754..3889
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3938..4732
                     /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             3938..4732
                     /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(3992..4011)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4602..4621
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4920..4939)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4964..5011
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5340..5928
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5829..5848
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag cacccagtcc aagctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggactcag atctcgagct caagcttcga attctgcagt cgagagggat
     1381 ctgggaagcg gaggaagcgg agaagaggaa gaaaagaaac aaaaagaaga acaagaaaga
     1441 cttgcaaaag aagaggcaga gagaaaacaa aaagaagaac aagaaagact tgcaaaagaa
     1501 gaggcagaga gaaaacaaaa ggaggaagaa gagagaaaac aaaaggaaga agaagagaga
     1561 aaacaaaagg aggaagaaga aagaaaatta aaggaggaac aagaaagaaa agctgcagaa
     1621 gaaaagaaag ctaaagaaga agctgagaga aaggctaaag aagaacaaga aaggaaagct
     1681 gaagaagaga gaaagaagaa agaagaggaa gaaagacttg aaagagaaag aaaagagaga
     1741 gaagaacaag aaaagaaagc caaagaagag gcagagagaa ttgcaaagtt agaggctgaa
     1801 aagaaggcag aagaagaaag aaaagccaaa gaagaagaag agagaaaagc caaagaagaa
     1861 gaggaaagaa agaagaaaga ggagcaagaa agacttgcaa aagaaaagga agaagcagaa
     1921 agaaaagctg cagaggaaaa gaaagctaaa gaagaacaag aaagaaaaga aaaggaagaa
     1981 gcagaaagaa aacaaagagg aagcggagga agcggaaagc ttggatccgg aatgtccaaa
     2041 ggagaagaac tgtttaccgg tgttgtgcca attttggttg aactcgatgg tgatgtcaac
     2101 ggacataagt tctcagtgag aggcgaagga gaaggtgacg ccaccattgg aaaattgact
     2161 cttaaattca tctgtactac tggtaaactt cctgtaccat ggccgactct cgtaacaacg
     2221 cttacgtacg gagttcagtg cttttcgaga tacccagacc atatgaaaag acatgacttt
     2281 tttaagtcgg ctatgcctga aggttacgtg caagaaagaa caatttcgtt caaagatgat
     2341 ggaaaatata aaactagagc agttgttaaa tttgaaggag atactttggt taaccgcatt
     2401 gaactgaaag gaacagattt taaagaagat ggtaatattc ttggacacaa actcgaatac
     2461 aattttaata gtcataacgt atacatcact gctgataagc aaaagaacgg aattaaagcg
     2521 aatttcacag tacgccataa tgtagaagat ggcagtgttc aacttgccga ccattaccaa
     2581 caaaacaccc ctattggaga cggtccggta cttcttcctg ataatcacta cctctcaaca
     2641 caaacagtcc tgagcaaaga tccaaatgaa aaaggaacag tcgacggtac cgcgggcccg
     2701 ggatccaccg gatctagata actgatcata atcagccata ccacatttgt agaggtttta
     2761 cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat gaatgcaatt
     2821 gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca
     2881 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc
     2941 aatgtatctt aaggcgtaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt
     3001 ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc
     3061 aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt
     3121 aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact
     3181 acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg
     3241 gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag
     3301 aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac
     3361 gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcaggtgg
     3421 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     3481 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     3541 gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc
     3601 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
     3661 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     3721 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     3781 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     3841 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     3901 aaagatcgat caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca
     3961 cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac
     4021 aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt
     4081 tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag cgcggctatc
     4141 gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg
     4201 aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc
     4261 tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc
     4321 ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat
     4381 ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc
     4441 cgaactgttc gccaggctca aggcgagcat gcccgacggc gaggatctcg tcgtgaccca
     4501 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga
     4561 ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat
     4621 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc
     4681 tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct gagcgggact
     4741 ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga tttcgattcc
     4801 accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg
     4861 atcctccagc gcggggatct catgctggag ttcttcgccc accctagggg gaggctaact
     4921 gaaacacgga aggagacaat accggaagga acccgcgcta tgacggcaat aaaaagacag
     4981 aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg
     5041 cactctgtcg ataccccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt
     5101 tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg
     5161 gcaggccctg ccatagcctc aggttactca tatatacttt agattgattt aaaacttcat
     5221 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     5281 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     5341 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     5401 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     5461 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     5521 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     5581 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     5641 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     5701 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     5761 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     5821 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     5881 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     5941 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     6001 ttatcccctg attctgtgga taaccgtatt accgccatgc at
//
