LOCUS       sequence-311033-        8135 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  tagging clonal populations with Peroxisomal acGFP fluorescent 
            protein. Adding Peroxisome localization signal to acGFP WC.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8135)
  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 8135)
  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: acGFP Peroxisome visual barcode
FEATURES             Location/Qualifiers
     source          1..8135
                     /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             1242..1958
                     /codon_start=1
                     /product="Aequorea coerulescens GFP"
                     /label=AcGFP1
                     /note="mammalian codon-optimized"
                     /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFEDD
                     GNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMIYF
                     GFVTAAAITHGMDELYK"
     misc_feature    1991..2579
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2044..2064)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2462..2473)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             2651..2884
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2962..3083
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2999..3018)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3053..3072
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      3123..3258
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3278..3297)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3279..3297)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3307..3324)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3307..3323)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3316..3338)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3465..3920
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3552..3571)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3762..3783
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3946..4050
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4051..4119,4120..4911)
                     /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(4269..4288)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5082..5670
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5571..5590
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5824..5841
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5958..5979
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5994..5999,6000..6017,6018..6024)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6032..6048
                     /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     6037..6059
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6056..6072
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6056..6072
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     6091..6111
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        6093..6111
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        6139..6365
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             6366..6546
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6593..6718
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7211..7444
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7629..7673
                     /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    7902..8019
                     /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 catggtgagc aagggcgccg
     1261 agctgttcac cggcatcgtg cccatcctga tcgagctgaa tggcgatgtg aatggccaca
     1321 agttcagcgt gagcggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt
     1381 tcatctgcac caccggcaag ctgcctgtgc cctggcccac cctggtgacc accctgagct
     1441 acggcgtgca gtgcttctca cgctaccccg atcacatgaa gcagcacgac ttcttcaaga
     1501 gcgccatgcc tgagggctac atccaggagc gcaccatctt cttcgaggat gacggcaact
     1561 acaagtcgcg cgccgaggtg aagttcgagg gcgataccct ggtgaatcgc atcgagctga
     1621 ccggcaccga tttcaaggag gatggcaaca tcctgggcaa taagatggag tacaactaca
     1681 acgcccacaa tgtgtacatc atgaccgaca aggccaagaa tggcatcaag gtgaacttca
     1741 agatccgcca caacatcgag gatggcagcg tgcagctggc cgaccactac cagcagaata
     1801 cccccatcgg cgatggccct gtgctgctgc ccgataacca ctacctgtcc acccagagcg
     1861 ccctgtccaa ggaccccaac gagaagcgcg atcacatgat ctacttcggc ttcgtgaccg
     1921 ccgccgccat cacccacggc atggatgagc tgtacaagtc ccgcctctaa taaagacgcg
     1981 ttaagtcgac aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa
     2041 ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat
     2101 tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta
     2161 tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc
     2221 aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt
     2281 ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg
     2341 ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc
     2401 ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc
     2461 ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct
     2521 tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgcg
     2581 tcgactttaa gaccaatgac ttacaaggca gctgtagatc ttagccactt tttaaaagaa
     2641 aaggggggac tggaagggct aattcactcc caacgaagac aagatctgct ttttgcttgt
     2701 actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac
     2761 ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg
     2821 ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct
     2881 agcagtacgt atagtagttc atgtcatctt attattcagt atttataact tgcaaagaaa
     2941 tgaatatcag agagtgagag gaacttgttt attgcagctt ataatggtta caaataaagc
     3001 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     3061 tccaaactca tcaatgtatc ttatcatgtc tggctctagc tatcccgccc ctaactccgc
     3121 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     3181 tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag
     3241 gaggcttttt tggaggccta gggacgtacc caattcgccc tatagtgagt cgtattacgc
     3301 gcgctcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact
     3361 taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac
     3421 cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tgggacgcgc cctgtagcgg
     3481 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     3541 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     3601 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     3661 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     3721 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     3781 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     3841 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa
     3901 aatattaacg cttacaattt aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     3961 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     4021 atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt
     4081 attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa
     4141 gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac
     4201 agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt
     4261 aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt
     4321 cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat
     4381 cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac
     4441 actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg
     4501 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
     4561 ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa
     4621 ctattaactg gcgaactact tactctagct tcccggcaac aattaataga ctggatggag
     4681 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
     4741 gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat
     4801 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     4861 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac
     4921 caagtttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     4981 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     5041 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     5101 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     5161 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     5221 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     5281 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     5341 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     5401 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     5461 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     5521 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     5581 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     5641 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     5701 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     5761 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     5821 cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc
     5881 gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc
     5941 agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac
     6001 tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga
     6061 aacagctatg accatgatta cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag
     6121 ctggagctgc aagcttaatg tagtcttatg caatactctt gtagtcttgc aacatggtaa
     6181 cgatgagtta gcaacatgcc ttacaaggag agaaaaagca ccgtgcatgc cgattggtgg
     6241 aagtaaggtg gtacgatcgt gccttattag gaaggcaaca gacgggtctg acatggattg
     6301 gacgaaccac tgaattgccg cattgcagag atattgtatt taagtgccta gctcgataca
     6361 taaacgggtc tctctggtta gaccagatct gagcctggga gctctctggc taactaggga
     6421 acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc
     6481 tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg tggaaaatct
     6541 ctagcagtgg cgcccgaaca gggacttgaa agcgaaaggg aaaccagagg agctctctcg
     6601 acgcaggact cggcttgctg aagcgcgcac ggcaagaggc gaggggcggc gactggtgag
     6661 tacgccaaaa attttgacta gcggaggcta gaaggagaga gatgggtgcg agagcgtcag
     6721 tattaagcgg gggagaatta gatcgcgatg ggaaaaaatt cggttaaggc cagggggaaa
     6781 gaaaaaatat aaattaaaac atatagtatg ggcaagcagg gagctagaac gattcgcagt
     6841 taatcctggc ctgttagaaa catcagaagg ctgtagacaa atactgggac agctacaacc
     6901 atcccttcag acaggatcag aagaacttag atcattatat aatacagtag caaccctcta
     6961 ttgtgtgcat caaaggatag agataaaaga caccaaggaa gctttagaca agatagagga
     7021 agagcaaaac aaaagtaaga ccaccgcaca gcaagcggcc gctgatcttc agacctggag
     7081 gaggagatat gagggacaat tggagaagtg aattatataa atataaagta gtaaaaattg
     7141 aaccattagg agtagcaccc accaaggcaa agagaagagt ggtgcagaga gaaaaaagag
     7201 cagtgggaat aggagctttg ttccttgggt tcttgggagc agcaggaagc actatgggcg
     7261 cagcgtcaat gacgctgacg gtacaggcca gacaattatt gtctggtata gtgcagcagc
     7321 agaacaattt gctgagggct attgaggcgc aacagcatct gttgcaactc acagtctggg
     7381 gcatcaagca gctccaggca agaatcctgg ctgtggaaag atacctaaag gatcaacagc
     7441 tcctggggat ttggggttgc tctggaaaac tcatttgcac cactgctgtg ccttggaatg
     7501 ctagttggag taataaatct ctggaacaga tttggaatca cacgacctgg atggagtggg
     7561 acagagaaat taacaattac acaagcttaa tacactcctt aattgaagaa tcgcaaaacc
     7621 agcaagaaaa gaatgaacaa gaattattgg aattagataa atgggcaagt ttgtggaatt
     7681 ggtttaacat aacaaattgg ctgtggtata taaaattatt cataatgata gtaggaggct
     7741 tggtaggttt aagaatagtt tttgctgtac tttctatagt gaatagagtt aggcagggat
     7801 attcaccatt atcgtttcag acccacctcc caaccccgag gggacccgat atcgcggccg
     7861 caattctaga tcttgagaca aatggcagta ttcatccaca attttaaaag aaaagggggg
     7921 attggggggt acagtgcagg ggaaagaata gtagacataa tagcaacaga catacaaact
     7981 aaagaattac aaaaacaaat tacaaaaatt caaaattttc gggtttatta cagggacagc
     8041 agagatccac tttggcgccg gctcgagggg gcccggggtt attaatagta atcaattacg
     8101 gggtcattag ttcatagccc atatatggag ttccg
//
