LOCUS       sequence-100661-        5577 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  5nm 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 5577)
  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 5577)
  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)-5nmSAH-GFP(1-10)
FEATURES             Location/Qualifiers
     source          1..5577
                     /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             1567..2208
                     /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    2365..2486
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2402..2421)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2456..2475
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2493..2948)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2630..2651)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2842..2861
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2975..3079
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3081..3438
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3081..3272
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3104..3124)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3289..3424
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3473..4267
                     /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             3473..4267
                     /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(3527..3546)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4137..4156
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4455..4474)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4499..4546
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4875..5463
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5364..5383
                     /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 ggagaggagg aggagcgcca gaagcgcgag gaggaggagc gcatcaagcg cctggaggag
     1501 ctggaggcga agcgcaagga ggaggagcgc aagggaagcg gaggaagcgg aaagcttgga
     1561 tccggaatgt ccaaaggaga agaactgttt accggtgttg tgccaatttt ggttgaactc
     1621 gatggtgatg tcaacggaca taagttctca gtgagaggcg aaggagaagg tgacgccacc
     1681 attggaaaat tgactcttaa attcatctgt actactggta aacttcctgt accatggccg
     1741 actctcgtaa caacgcttac gtacggagtt cagtgctttt cgagataccc agaccatatg
     1801 aaaagacatg acttttttaa gtcggctatg cctgaaggtt acgtgcaaga aagaacaatt
     1861 tcgttcaaag atgatggaaa atataaaact agagcagttg ttaaatttga aggagatact
     1921 ttggttaacc gcattgaact gaaaggaaca gattttaaag aagatggtaa tattcttgga
     1981 cacaaactcg aatacaattt taatagtcat aacgtataca tcactgctga taagcaaaag
     2041 aacggaatta aagcgaattt cacagtacgc cataatgtag aagatggcag tgttcaactt
     2101 gccgaccatt accaacaaaa cacccctatt ggagacggtc cggtacttct tcctgataat
     2161 cactacctct caacacaaac agtcctgagc aaagatccaa atgaaaaagg aacagtcgac
     2221 ggtaccgcgg gcccgggatc caccggatct agataactga tcataatcag ccataccaca
     2281 tttgtagagg ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat
     2341 aaaatgaatg caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa
     2401 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     2461 ttgtccaaac tcatcaatgt atcttaaggc gtaaattgta agcgttaata ttttgttaaa
     2521 attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa
     2581 aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa
     2641 caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     2701 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     2761 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     2821 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     2881 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     2941 gggcgcgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     3001 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     3061 atattgaaaa aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta
     3121 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     3181 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3241 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     3301 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     3361 gaggccgagg ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     3421 ggcctaggct tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga
     3481 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
     3541 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
     3601 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga
     3661 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
     3721 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
     3781 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
     3841 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
     3901 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
     3961 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga
     4021 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
     4081 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
     4141 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
     4201 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
     4261 cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca
     4321 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg
     4381 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct
     4441 agggggaggc taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg
     4501 gcaataaaaa gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt
     4561 cggtcccagg gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc
     4621 cgcgtttctt ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca
     4681 gccaacgtcg gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt
     4741 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     4801 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     4861 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     4921 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     4981 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
     5041 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     5101 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     5161 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     5221 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     5281 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     5341 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     5401 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     5461 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     5521 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcat
//
