LOCUS       sequence-386306-        5386 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 0 PhiC31 inegrase NLS with GG2 GG4 Golden gate linkers.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5386)
  AUTHORS   Guiziou S, Maranas CJ, Chu JC, Nemhauser JL
  TITLE     An integrase toolbox to record gene-expression during plant 
            development.
  JOURNAL   Nat Commun. 2023 Apr 3;14(1):1844. doi: 10.1038/s41467-023-37607-5.
  PUBMED    37012288
REFERENCE   2  (bases 1 to 5386)
  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: "2023-04-3"; volume: "14(1):1844. doi"; pages: " 
            10.1038/s41467-023-37607-5"
COMMENT     Sequence Label: L0-I12 PhiC31-NLS GG2-GG4
FEATURES             Location/Qualifiers
     source          1..5386
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             20..1831
                     /codon_start=1
                     /product="integrase from phage phi-C31"
                     /label=phage phi-C31 integrase
                     /translation="DTYAGAYDRQSRERENSSAASPATQRSANEDKAADLQREVERDGG
                     RFRFVGHFSEAPGTSAFGTAERPEFERILNECRAGRLNMIIVYDVSRFSRLKVMDAIPI
                     VSELLALGVTIVSTQEGVFRQGNVMDLIHLIMRLDASHKESSLKSAKILDTKNLQRELG
                     GYVGGKAPYGFELVSETKEITRNGRMVNVVINKLAHSTTPLTGPFEFEPDVIRWWWREI
                     KTHKHLPFKPGSQAAIHPGSITGLCKRMDADAVPTRGETIGKKTASSAWDPATVMRILR
                     DPRIAGFAAEVIYKKKPDGTPTTKIEGYRIQRDPITLRPVELDCGPIIEPAEWYELQAW
                     LDGRGRGKGLSRGQAILSAMDKLYCECGAVMTSKRGEESIKDSYRCRRRKVVDPSAPGQ
                     HEGTCNVSMAALDKFVAERIFNKIRHAEGDEETLALLWEAARRFGKLTEAPEKSGERAN
                     LVAERADALNALEELYEDRAAGAYDGPVGRKHFRKQQAALTLRQQGAEERLAELEAAEA
                     PKLPLDQWFPEDADADPTGPKSWWGRASVDDKRVFVGLFVDKIVVTKSTTGRGQGTPIE
                     KRASITWAKPPTDDDEDDAQDGTEDVAA"
     CDS             1832..1852
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(1987..2003)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1987..2003)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2000..2022)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2011..2027
                     /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(2035..2041,2042..2059,2060..2065))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2080..2101
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2218..2235)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2389..2977)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2469..2488)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3115..3489)
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     regulatory      3486..3495
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(3695..4489)
                     /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             complement(3695..4489)
                     /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(3806..3825)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4416..4435
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     CDS             complement(4838..5140)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(4940..4959)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     primer_bind     5263..5285
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5277..5294
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5278..5294
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5301..5320
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5301..5319
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 cactctgtgg tctcaagccg acacgtatgc cggcgcttat gatagacaat caagagaaag
       61 agagaactct tctgccgcct ctccagctac acagagatct gccaacgagg ataaagctgc
      121 tgatttacaa agagaggtag agagagatgg gggtaggttt agatttgtgg gtcatttctc
      181 tgaggctcct ggtacatcag cctttggtac tgctgaacgg cctgaatttg aaagaatatt
      241 aaatgagtgt cgggccggtc gactaaatat gataatagtt tatgatgtat ctaggttcag
      301 tagactcaaa gttatggatg caattccgat cgtgagcgag ctacttgcac tcggtgttac
      361 tatcgtaagt acacaagaag gtgtattcag gcagggtaat gtgatggacc ttatacacct
      421 tattatgcga ctcgacgcta gccataaaga gtcatcactg aagagtgcta aaatcttgga
      481 caccaagaac ctgcaaaggg agcttggagg ctatgtagga ggaaaagctc cgtatggttt
      541 cgaactcgtg agtgagacaa aagagattac cagaaatggt aggatggtga acgtggttat
      601 taataagcta gctcatagca caacccctct cacagggcct tttgagttcg aacctgacgt
      661 tattagatgg tggtggagag agatcaaaac tcacaaacat cttccattta agccaggtag
      721 ccaggctgct atccaccctg gatctatcac tgggctttgt aaaaggatgg atgctgacgc
      781 agtgccaact cgaggggaaa caatcggtaa gaagactgca tcttccgctt gggatcccgc
      841 tacagttatg agaatccttc gggatccgcg tattgctgga ttcgccgcag aagtcattta
      901 taagaaaaaa cctgatggta ctcctactac aaaaatcgaa ggttatagga tccaaaggga
      961 tcctattact ttgcgtccag ttgaattaga ctgtggaccg atcatcgaac ctgccgagtg
     1021 gtacgagttg caggcctggc ttgatggtag agggcgtgga aagggtttat ctcggggaca
     1081 ggcaattttg tcagcaatgg ataagcttta ttgtgagtgt ggggctgtta tgacatcaaa
     1141 acgtggagaa gagtcaatca aggattctta caggtgcaga cgtcgtaaag tggtggaccc
     1201 ttctgctcca ggacagcatg agggtacatg caacgtttct atggctgctc ttgataaatt
     1261 tgtggccgaa cgtattttca acaagattag acacgctgaa ggggatgagg aaactttagc
     1321 tcttctttgg gaggctgccc ggcgttttgg taagctaaca gaggcaccag aaaaaagtgg
     1381 cgaaagagct aatttggtag ccgaaagagc agacgccttg aatgcattgg aagaacttta
     1441 cgaagatagg gctgccggag catatgatgg accagttggt cggaagcatt tcaggaagca
     1501 acaagctgca ctaactttga gacagcaagg cgcagaggag cgactcgcag agcttgaggc
     1561 cgccgaggcc ccaaagcttc cactcgatca atggtttcct gaggatgccg acgctgatcc
     1621 tacaggtcca aagagctggt ggggaagagc tagcgtagac gataagagag tgtttgtggg
     1681 tctatttgtt gacaagattg tagttactaa gtccactacg ggtcgaggac agggcactcc
     1741 catcgaaaag agagccagta ttacctgggc aaagcctcca accgatgatg atgaagacga
     1801 tgcacaagat ggtaccgagg atgtggcagc accaaaaaaa aagaggaagg tgtaagcttt
     1861 gagacctcat cacgcctgaa ttccagcaca ctggcggccg ttactagtgg atccgagctc
     1921 ggtaccaagc ttgatgcata gcttgagtat tctaacgcgt cacctaaata gcttggcgta
     1981 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     2041 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     2101 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     2161 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     2221 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     2281 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     2341 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     2401 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     2461 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     2521 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     2581 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     2641 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     2701 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     2761 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     2821 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     2881 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     2941 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     3001 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     3061 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttagca cgtgtcagtc
     3121 ctgctcctcg gccacgaagt gcacgcagtt gccggccggg tcgcgcaggg cgaactcccg
     3181 cccccacggc tgctcgccga tctcggtcat ggccggcccg gaggcgtccc ggaagttcgt
     3241 ggacacgacc tccgaccact cggcgtacag ctcgtccagg ccgcgcaccc acacccaggc
     3301 cagggtgttg tccggcacca cctggtcctg gaccgcgctg atgaacaggg tcacgtcgtc
     3361 ccggaccaca ccggcgaagt cgtcctccac gaagtcccgg gagaacccga gccggtcggt
     3421 ccagaactcg accgctccgg cgacgtcgcg cgcggtgagc accggaacgg cactggtcaa
     3481 cttggccatg gtggccctcc tcacgtgcta ttattgaagc atttatcagg gttattgtct
     3541 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     3601 atttccccga aaagtgccac ctgatgcggt gtgaaatacc gcacagatgc gtaaggagaa
     3661 aataccgcat caggaaattg taagcgttaa taattcagaa gaactcgtca agaaggcgat
     3721 agaaggcgat gcgctgcgaa tcgggagcgg cgataccgta aagcacgagg aagcggtcag
     3781 cccattcgcc gccaagctct tcagcaatat cacgggtagc caacgctatg tcctgatagc
     3841 ggtccgccac acccagccgg ccacagtcga tgaatccaga aaagcggcca ttttccacca
     3901 tgatattcgg caagcaggca tcgccatggg tcacgacgag atcctcgccg tcgggcatgc
     3961 tcgccttgag cctggcgaac agttcggctg gcgcgagccc ctgatgctct tcgtccagat
     4021 catcctgatc gacaagaccg gcttccatcc gagtacgtgc tcgctcgatg cgatgtttcg
     4081 cttggtggtc gaatgggcag gtagccggat caagcgtatg cagccgccgc attgcatcag
     4141 ccatgatgga tactttctcg gcaggagcaa ggtgagatga caggagatcc tgccccggca
     4201 cttcgcccaa tagcagccag tcccttcccg cttcagtgac aacgtcgagc acagctgcgc
     4261 aaggaacgcc cgtcgtggcc agccacgata gccgcgctgc ctcgtcttgc agttcattca
     4321 gggcaccgga caggtcggtc ttgacaaaaa gaaccgggcg cccctgcgct gacagccgga
     4381 acacggcggc atcagagcag ccgattgtct gttgtgccca gtcatagccg aatagcctct
     4441 ccacccaagc ggccggagaa cctgcgtgca atccatcttg ttcaatcatg cgaaacgatc
     4501 ctcatcctgt ctcttgatca gagcttgatc ccctgcgcca tcagatcctt ggcggcaaga
     4561 aagccatcca gtttactttg cagggcttcc caaccttacc agagggcgcc ccagctggca
     4621 attccggttc gcttgctgtc cataaaaccg cccagtctag ctatcgccat gtaagcccac
     4681 tgcaagctac ctgctttctc tttgcgcttg cgttttccct tgtccagata gcccagtagc
     4741 tgacattcat ccggggtcag caccgtttct gcggactggc tttctacgtg aaaaggatct
     4801 aggtgaagat cctttttgat aatctcatgc ctgacattta tattccccag aacatcaggt
     4861 taatggcgtt tttgatgtca ttttcgcggt ggctgagatc agccacttct tccccgataa
     4921 cggagaccgg cacactggcc atatcggtgg tcatcatgcg ccagctttca tccccgatat
     4981 gcaccaccgg gtaaagttca cgggagactt tatctgacag cagacgtgca ctggccaggg
     5041 ggatcaccat ccgtcgcccc ggcgtgtcaa taatatcact ctgtacatcc acaaacagac
     5101 gataacggct ctctctttta taggtgtaaa ccttaaactg ccgtacgtat aggctgcgca
     5161 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg
     5221 gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta
     5281 aaacgacggc cagtgaattg taatacgact cactataggg cgaattgggc cctctagatg
     5341 catgctcgag cggccgccag tgtgatggat atctgcagaa ttcagg
//
