LOCUS       sequence-311022-        8189 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  tagging clonal populations with NES mTAGBFP2 fluorescent protein. 
            Adding NES to mTAGBFP2 WC.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8189)
  AUTHORS   Kaufman T, Nitzan E, Firestein N, Ginzberg MB, Iyengar S, Patel N, 
            Ben-Hamo R, Porat Z, Hunter J, Hilfinger A, Rotter V, Kafri R, 
            Straussman R
  TITLE     Visual barcodes for clonal-multiplexing of live microscopy-based 
            assays.
  JOURNAL   Nat Commun. 2022 May 18;13(1):2725. doi: 10.1038/s41467-022-30008-0.
  PUBMED    35585055
REFERENCE   2  (bases 1 to 8189)
  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: "Nat 
            Commun."; date: "2022-05-18"; volume: "13(1):2725. doi"; pages: " 
            10.1038/s41467-022-30008-0"
COMMENT     Sequence Label: mTAGBFP2 NES visual barcode
FEATURES             Location/Qualifiers
     source          1..8189
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     complement(534..553)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1030..1050
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             1170..1235
                     /codon_start=1
                     /product="2A peptide from foot-and-mouth disease virus 
                     polyprotein"
                     /label=F2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="VKQTLNFDLLKLAGDVESNPGP"
     CDS             1251..1268
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1278..1976
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     misc_feature    2045..2633
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2098..2118)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2516..2527)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             2705..2938
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    3016..3137
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3053..3072)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3107..3126
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      3177..3312
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3332..3351)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3333..3351)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3361..3378)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3361..3377)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3370..3392)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3519..3974
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3606..3625)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3816..3837
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4000..4104
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4105..4173,4174..4965)
                     /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     complement(4323..4342)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5136..5724
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5625..5644
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5878..5895
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6012..6033
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6048..6053,6054..6071,6072..6078)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6086..6102
                     /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)."
     primer_bind     6091..6113
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6110..6126
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6110..6126
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6145..6165
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        6147..6165
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        6193..6419
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             6420..6600
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6647..6772
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7265..7498
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7683..7727
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    7956..8073
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctct ctggctaact gtcgggatcc accatgaccg
      541 agtacaagcc caccgtgcgc ctcgccaccc gcgacgacgt gcccagggcc gtccgcaccc
      601 tcgccgccgc tttcgccgac taccccgcca cccgccacac cgtcgatccc gaccgccaca
      661 tcgagcgggt caccgagctg caagaactct tcctcacccg cgtcgggctc gacatcggca
      721 aggtgtgggt cgctgacgac ggcgccgcag tggctgtctg gaccacaccc gagagcgtcg
      781 aagctggggc ggtgttcgcc gagatcggcc ctcgcatggc cgagttgagc ggttcccggc
      841 tggccgctca gcaacagatg gaaggcctcc tggctccaca ccggcccaag gagcccgcat
      901 ggttcctggc caccgtcggc gtctctcccg accaccaggg caagggtctg ggcagcgccg
      961 tcgtgctccc cggagtggag gctgccgagc gcgccggggt gcccgccttc ctggagacct
     1021 ccgcaccccg caacctcccc ttctacgagc ggctcggctt caccgtcacc gccgacgtgg
     1081 aggtgcccga aggaccacgc acctggtgca tgacccgcaa gcccggtgcc ctgcagggat
     1141 ctggcgaagc cagacacaaa cagaaaattg tgaaacagac tttgaatttt gaccttctca
     1201 agttggcggg agacgtggag tccaacccag ggcccgctag catggggagc caccatcacc
     1261 atcaccatgg cagatctatg agcgagctga ttaaggagaa catgcacatg aagctgtaca
     1321 tggagggcac cgtggacaac catcacttca agtgcacatc cgagggcgaa ggcaagccct
     1381 acgagggcac ccagaccatg agaatcaagg tggtcgaggg cggccctctc cccttcgcct
     1441 tcgacatcct ggctactagc ttcctctacg gcagcaagac cttcatcaac cacacccagg
     1501 gcatccccga cttcttcaag cagtccttcc ctgagggctt cacatgggag agagtcacca
     1561 catacgaaga cgggggcgtg ctgaccgcta cccaggacac cagcctccag gacggctgcc
     1621 tcatctacaa cgtcaagatc agaggggtga acttcacatc caacggccct gtgatgcaga
     1681 agaaaacact cggctgggag gccttcaccg agacgctgta ccccgctgac ggcggcctgg
     1741 aaggcagaaa cgacatggcc ctgaagctcg tgggcgggag ccatctgatc gcaaacgcca
     1801 agaccacata tagatccaag aaacccgcta agaacctcaa gatgcctggc gtctactatg
     1861 tggactacag actggaaaga atcaaggagg ccaacaacga gacctacgtc gagcagcacg
     1921 aggtggcagt ggccagatac tgcgacctcc ctagcaaact ggggcacaag cttaatagtg
     1981 agctgcagaa caagctggaa gagttggatc tggactcgta caagtaaaga cgcgttaagt
     2041 cgacaatcaa cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt
     2101 tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc
     2161 ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga
     2221 gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc
     2281 cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct
     2341 ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg
     2401 gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct
     2461 gctcgcctgt gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc
     2521 cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg
     2581 tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcgtcgact
     2641 ttaagaccaa tgacttacaa ggcagctgta gatcttagcc actttttaaa agaaaagggg
     2701 ggactggaag ggctaattca ctcccaacga agacaagatc tgctttttgc ttgtactggg
     2761 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
     2821 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
     2881 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt
     2941 acgtatagta gttcatgtca tcttattatt cagtatttat aacttgcaaa gaaatgaata
     3001 tcagagagtg agaggaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     3061 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     3121 ctcatcaatg tatcttatca tgtctggctc tagctatccc gcccctaact ccgcccatcc
     3181 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     3241 tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct
     3301 tttttggagg cctagggacg tacccaattc gccctatagt gagtcgtatt acgcgcgctc
     3361 actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg
     3421 ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg
     3481 cccttcccaa cagttgcgca gcctgaatgg cgaatgggac gcgccctgta gcggcgcatt
     3541 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     3601 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     3661 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     3721 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     3781 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     3841 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     3901 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt
     3961 aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     4021 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     4081 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     4141 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     4201 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     4261 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     4321 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     4381 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     4441 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     4501 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     4561 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     4621 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     4681 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     4741 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     4801 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     4861 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     4921 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     4981 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     5041 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     5101 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     5161 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     5221 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     5281 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     5341 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     5401 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     5461 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     5521 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     5581 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     5641 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     5701 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     5761 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     5821 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     5881 gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt
     5941 tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag
     6001 cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg
     6061 cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc
     6121 tatgaccatg attacgccaa gcgcgcaatt aaccctcact aaagggaaca aaagctggag
     6181 ctgcaagctt aatgtagtct tatgcaatac tcttgtagtc ttgcaacatg gtaacgatga
     6241 gttagcaaca tgccttacaa ggagagaaaa agcaccgtgc atgccgattg gtggaagtaa
     6301 ggtggtacga tcgtgcctta ttaggaaggc aacagacggg tctgacatgg attggacgaa
     6361 ccactgaatt gccgcattgc agagatattg tatttaagtg cctagctcga tacataaacg
     6421 ggtctctctg gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac
     6481 tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt
     6541 gtgactctgg taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca
     6601 gtggcgcccg aacagggact tgaaagcgaa agggaaacca gaggagctct ctcgacgcag
     6661 gactcggctt gctgaagcgc gcacggcaag aggcgagggg cggcgactgg tgagtacgcc
     6721 aaaaattttg actagcggag gctagaagga gagagatggg tgcgagagcg tcagtattaa
     6781 gcgggggaga attagatcgc gatgggaaaa aattcggtta aggccagggg gaaagaaaaa
     6841 atataaatta aaacatatag tatgggcaag cagggagcta gaacgattcg cagttaatcc
     6901 tggcctgtta gaaacatcag aaggctgtag acaaatactg ggacagctac aaccatccct
     6961 tcagacagga tcagaagaac ttagatcatt atataataca gtagcaaccc tctattgtgt
     7021 gcatcaaagg atagagataa aagacaccaa ggaagcttta gacaagatag aggaagagca
     7081 aaacaaaagt aagaccaccg cacagcaagc ggccgctgat cttcagacct ggaggaggag
     7141 atatgaggga caattggaga agtgaattat ataaatataa agtagtaaaa attgaaccat
     7201 taggagtagc acccaccaag gcaaagagaa gagtggtgca gagagaaaaa agagcagtgg
     7261 gaataggagc tttgttcctt gggttcttgg gagcagcagg aagcactatg ggcgcagcgt
     7321 caatgacgct gacggtacag gccagacaat tattgtctgg tatagtgcag cagcagaaca
     7381 atttgctgag ggctattgag gcgcaacagc atctgttgca actcacagtc tggggcatca
     7441 agcagctcca ggcaagaatc ctggctgtgg aaagatacct aaaggatcaa cagctcctgg
     7501 ggatttgggg ttgctctgga aaactcattt gcaccactgc tgtgccttgg aatgctagtt
     7561 ggagtaataa atctctggaa cagatttgga atcacacgac ctggatggag tgggacagag
     7621 aaattaacaa ttacacaagc ttaatacact ccttaattga agaatcgcaa aaccagcaag
     7681 aaaagaatga acaagaatta ttggaattag ataaatgggc aagtttgtgg aattggttta
     7741 acataacaaa ttggctgtgg tatataaaat tattcataat gatagtagga ggcttggtag
     7801 gtttaagaat agtttttgct gtactttcta tagtgaatag agttaggcag ggatattcac
     7861 cattatcgtt tcagacccac ctcccaaccc cgaggggacc cgatatcgcg gccgcaattc
     7921 tagatcttga gacaaatggc agtattcatc cacaatttta aaagaaaagg ggggattggg
     7981 gggtacagtg caggggaaag aatagtagac ataatagcaa cagacataca aactaaagaa
     8041 ttacaaaaac aaattacaaa aattcaaaat tttcgggttt attacaggga cagcagagat
     8101 ccactttggc gccggctcga gggggcccgg ggttattaat agtaatcaat tacggggtca
     8161 ttagttcata gcccatatat ggagttccg
//
