LOCUS       sequence-320862-        6077 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GFP(9-7) in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6077)
  AUTHORS   Tamura R, Jiang F, Xie J, Kamiyama D
  TITLE     Multiplexed labeling of cellular proteins with split fluorescent 
            protein tags.
  JOURNAL   Commun Biol. 2021 Feb 26;4(1):257. doi: 10.1038/s42003-021-01780-4.
  PUBMED    33637968
REFERENCE   2  (bases 1 to 6077)
  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"; journalName: "Commun 
            Biol."; date: "2021-02-26"; volume: "4(1):257. doi"; pages: " 
            10.1038/s42003-021-01780-4"
COMMENT     Sequence Label: pcDNA3.1-GFP(9-7)
FEATURES             Location/Qualifiers
     source          1..6077
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1061..1108
                     /codon_start=1
                     /product="11th beta-strand of superfolder GFP"
                     /label=GFP11
                     /note="associates with GFP(1-10), consisting of the first 
                     10 beta-strands, to reconstitute fluorescent GFP (Cabantous
                     et al., 2005; Kamiyama et al., 2016)"
                     /translation="RDHMVLHEYVNAAGIT"
     primer_bind     complement(1671..1688)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1677..1901
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1947..2375
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2034..2053)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2244..2265
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2384..2404)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2389..2718
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2569..2704
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2631..2650
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2785..3579
                     /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(2839..2858)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3449..3468
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3753..3874
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3790..3809)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3844..3863
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3923..3939)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3923..3939)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3936..3958)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3947..3963
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(3971..3977,3978..3995,3996..4001))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4016..4037
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4154..4171)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4325..4910)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4405..4424)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5081..5872,5873..5941))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5704..5723
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5942..6046)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttggta ccggatccat ggaagatggc agtgttcaac ttgccgacca
      961 ttaccaacaa aacaccccta ttggagacgg tccggtactt cttcctgata atcactacct
     1021 ctcaacacaa acagtcctga gcaaagatcc aaatgaaaaa cgtgaccaca tggtccttca
     1081 tgagtatgta aatgctgctg ggattacagg tggcaccggt ggctctatgt ccaaaggaga
     1141 agaactgttt accggtgttg tgccaatttt ggttgaactc gatggtgatg tcaacggaca
     1201 taagttctca gtgagaggcg aaggagaagg tgacgccacc attggaaaat tgactcttaa
     1261 attcatctgt actactggta aacttcctgt accatggccg actctcgtaa caacgcttac
     1321 gtacggagtt cagtgctttt cgagataccc agaccatatg aaaagacatg acttttttaa
     1381 gtcggctatg cctgaaggtt acgtgcaaga aagaacaatt tcgttcaaag atgatggaaa
     1441 atataaaact agagcagttg ttaaatttga aggagatact ttggttaacc gcattgaact
     1501 gaaaggaaca gattttaaag aagatggtaa tattcttgga cacaaactcg aatacaattt
     1561 taatagtcat aacgtataca tcactgctga taagcaatag gaattctgca gatatccagc
     1621 acagtggcgg ccgctcgagt ctagagggcc cgtttaaacc cgctgatcag cctcgactgt
     1681 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     1741 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     1801 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     1861 agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     1921 cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg
     1981 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     2041 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     2101 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     2161 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     2221 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     2281 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     2341 aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta
     2401 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
     2461 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     2521 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
     2581 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
     2641 gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga
     2701 ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcaa
     2761 gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg
     2821 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     2881 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     2941 ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg
     3001 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     3061 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     3121 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     3181 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     3241 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     3301 aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     3361 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     3421 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     3481 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     3541 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa
     3601 tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct
     3661 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     3721 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt
     3781 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     3841 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta
     3901 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     3961 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     4021 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     4081 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     4141 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     4201 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     4261 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     4321 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     4381 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     4441 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     4501 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     4561 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     4621 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     4681 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     4741 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     4801 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     4861 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     4921 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     4981 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     5041 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     5101 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     5161 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     5221 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     5281 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     5341 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     5401 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     5461 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     5521 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     5581 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     5641 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     5701 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     5761 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     5821 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     5881 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     5941 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     6001 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     6061 aagtgccacc tgacgtc
//
