LOCUS       sequence-100657-        6732 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  2x-30nm SAH scaffold with EF Hand and split GFP attachment sites.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6732)
  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 6732)
  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: SNAP-EF(N)-60nmSAH-GFP(1-10)
FEATURES             Location/Qualifiers
     source          1..6732
                     /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             2722..3363
                     /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    3520..3641
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3557..3576)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3611..3630
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3648..4103)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3785..3806)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3997..4016
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4130..4234
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4236..4593
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4236..4427
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4259..4279)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4444..4579
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4628..5422
                     /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             4628..5422
                     /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(4682..4701)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5292..5311
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5610..5629)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5654..5701
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6030..6618
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6519..6538
                     /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 ccatggacaa ggattgtgaa atgaaacgca ccacactgga cagccctttg
      661 gggaagctgg agctgtctgg ttgtgagcag ggtctgcacg aaataaagct cctgggcaag
      721 gggacgtctg cagctgatgc cgtggaggtc ccagcccccg ctgcggttct cggaggtccg
      781 gagcccctga tgcaatgcac agcctggctg aatgcctatt tccaccagcc cgaggctatc
      841 gaagagttcc ccgtgccggc tcttcaccat cccgttttcc agcaagagtc gttcaccaga
      901 caggtgttat ggaagctgct gaaggttgtg aaattcggag aagtgatttc ttaccagcaa
      961 ttagcagccc tggcaggcaa ccccgcagcc acggcagcag tgggaggagc aatgagaggc
     1021 aatcctgtcc ctatcctgat cccgtgccac agagtggtct gcagcagcgg agccgtgggc
     1081 ggttacgagg gtggactggc cgtgaaggaa tggcttctgg cccatgaagg ccatcggttg
     1141 gggaagccag gcttgggagg gagctcaggt ctggcagggg cctggctcaa gggagcggga
     1201 gctacctcgg gctccccgcc tgctggccga aacgtgtaca agtccggaag cggtggatca
     1261 ggcgctagca tgaaatcccc ggaagaactc aaaggaatct ttgaaaagta cgccgcgaag
     1321 gagggggacc ccaaccaatt gtcgaaggag gagcttaaac tgttgcttca gacggagttc
     1381 ccttcgctgc tcaaggggat gggttcgggg ctcgagaggg atctgggaag cggaggaagc
     1441 ggagaagagg aagaaaagaa acaaaaagaa gaacaagaaa gacttgcaaa agaagaggca
     1501 gagagaaaac aaaaagaaga acaagaaaga cttgcaaaag aagaggcaga gagaaaacaa
     1561 aaggaggaag aagagagaaa acaaaaggaa gaagaagaga gaaaacaaaa ggaggaagaa
     1621 gaaagaaaat taaaggagga acaagaaaga aaagctgcag aagaaaagaa agctaaagaa
     1681 gaagctgaga gaaaggctaa agaagaacaa gaaaggaaag ctgaagaaga gagaaagaag
     1741 aaagaagagg aagaaagact tgaaagagaa agaaaagaga gagaagaaca agaaaagaaa
     1801 gccaaagaag aggcagagag aattgcaaag ttagaggctg aaaagaaggc agaagaagaa
     1861 agaaaagcca aagaagaaga agagagaaaa gccaaagaag aagaggaaag aaagaagaaa
     1921 gaggagcaag aaagacttgc aaaagaaaag gaagaagcag aaagaaaagc tgcagaggaa
     1981 aagaaagcta aagaagaaca agaaagaaaa gaaaaggaag aagcagaaag aaaacaaaga
     2041 ggaagcggag gaagcggaaa gctgcgggat ctgggaagcg gaggaagcgg agaagaggaa
     2101 gaaaagaaac aaaaagaaga acaagaaaga cttgcaaaag aagaggcaga gagaaaacaa
     2161 aaagaagaac aagaaagact tgcaaaagaa gaggcagaga gaaaacaaaa ggaggaagaa
     2221 gagagaaaac aaaaggaaga agaagagaga aaacaaaagg aggaagaaga aagaaaatta
     2281 aaggaggaac aagaaagaaa agctgcagaa gaaaagaaag ctaaagaaga agctgagaga
     2341 aaggctaaag aagaacaaga aaggaaagct gaagaagaga gaaagaagaa agaagaggaa
     2401 gaaagacttg aaagagaaag aaaagagaga gaagaacaag aaaagaaagc caaagaagag
     2461 gcagagagaa ttgcaaagtt agaggctgaa aagaaggcag aagaagaaag aaaagccaaa
     2521 gaagaagaag agagaaaagc caaagaagaa gaggaaagaa agaagaaaga ggagcaagaa
     2581 agacttgcaa aagaaaagga agaagcagaa agaaaagctg cagaggaaaa gaaagctaaa
     2641 gaagaacaag aaagaaaaga aaaggaagaa gcagaaagaa aacaaagagg aagcggagga
     2701 agcggaaagc ttggatccgg aatgtccaaa ggagaagaac tgtttaccgg tgttgtgcca
     2761 attttggttg aactcgatgg tgatgtcaac ggacataagt tctcagtgag aggcgaagga
     2821 gaaggtgacg ccaccattgg aaaattgact cttaaattca tctgtactac tggtaaactt
     2881 cctgtaccat ggccgactct cgtaacaacg cttacgtacg gagttcagtg cttttcgaga
     2941 tacccagacc atatgaaaag acatgacttt tttaagtcgg ctatgcctga aggttacgtg
     3001 caagaaagaa caatttcgtt caaagatgat ggaaaatata aaactagagc agttgttaaa
     3061 tttgaaggag atactttggt taaccgcatt gaactgaaag gaacagattt taaagaagat
     3121 ggtaatattc ttggacacaa actcgaatac aattttaata gtcataacgt atacatcact
     3181 gctgataagc aaaagaacgg aattaaagcg aatttcacag tacgccataa tgtagaagat
     3241 ggcagtgttc aacttgccga ccattaccaa caaaacaccc ctattggaga cggtccggta
     3301 cttcttcctg ataatcacta cctctcaaca caaacagtcc tgagcaaaga tccaaatgaa
     3361 aaaggaacag tcgacggtac cgcgggcccg ggatccaccg gatctagata actgatcata
     3421 atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc
     3481 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
     3541 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     3601 cattctagtt gtggtttgtc caaactcatc aatgtatctt aaggcgtaaa ttgtaagcgt
     3661 taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata
     3721 ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt
     3781 tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg
     3841 aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt
     3901 ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc
     3961 ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg
     4021 cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct
     4081 taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     4141 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     4201 ataaatgctt caataatatt gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg
     4261 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     4321 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     4381 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     4441 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     4501 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     4561 ggaggctttt ttggaggcct aggcttttgc aaagatcgat caagagacag gatgaggatc
     4621 gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     4681 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     4741 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     4801 tgaactgcaa gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     4861 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     4921 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     4981 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     5041 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     5101 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat
     5161 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     5221 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     5281 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     5341 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     5401 ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg
     5461 cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     5521 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     5581 ttcttcgccc accctagggg gaggctaact gaaacacgga aggagacaat accggaagga
     5641 acccgcgcta tgacggcaat aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt
     5701 cataaacgcg gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat
     5761 tggggccaat acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa
     5821 ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc aggttactca
     5881 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     5941 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     6001 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     6061 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     6121 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt
     6181 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     6241 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     6301 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     6361 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     6421 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     6481 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     6541 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     6601 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     6661 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     6721 accgccatgc at
//
