LOCUS       sequence-386344-        4564 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 0 Intein(DnaE-Ssp)-PhiC31 with GG1 and GG4 golden gate 
            linkers.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4564)
  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 4564)
  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-I40 Intein-PhiC31 GG1-GG4
FEATURES             Location/Qualifiers
     source          1..4564
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1010..1030
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(1165..1181)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1165..1181)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1178..1200)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1189..1205
                     /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(1213..1219,1220..1237,1238..1243))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1258..1279
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1396..1413)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1567..2155)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1647..1666)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2293..2667)
                     /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      2664..2673
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(2873..3667)
                     /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(2873..3667)
                     /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(2984..3003)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3594..3613
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     CDS             complement(4016..4318)
                     /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(4118..4137)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     primer_bind     4441..4463
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4455..4472
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4456..4472
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4479..4498
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4479..4497
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggatcggagg tctcaaatgg tgaaagtgat tggccgccgc agcctgggcg tgcagcgcat
       61 ttttgatatt ggcctgcgcc aggatcataa ctttctgctg gcgaacggcg cgattgcggc
      121 gaactgcttt aacagggatc ctattacttt gcgtccagtt gaattagact gtggaccgat
      181 catcgaacct gccgagtggt acgagttgca ggcctggctt gatggtagag ggcgtggaaa
      241 gggtttatct cggggacagg caattttgtc agcaatggat aagctttatt gtgagtgtgg
      301 ggctgttatg acatcaaaac gtggagaaga gtcaatcaag gattcttaca ggtgcagacg
      361 tcgtaaagtg gtggaccctt ctgctccagg acagcatgag ggtacatgca acgtttctat
      421 ggctgctctt gataaatttg tggccgaacg tattttcaac aagattagac acgctgaagg
      481 ggatgaggaa actttagctc ttctttggga ggctgcccgg cgttttggta agctaacaga
      541 ggcaccagaa aaaagtggcg aaagagctaa tttggtagcc gaaagagcag acgccttgaa
      601 tgcattggaa gaactttacg aagatagggc tgccggagca tatgatggac cagttggtcg
      661 gaagcatttc aggaagcaac aagctgcact aactttgaga cagcaaggcg cagaggagcg
      721 actcgcagag cttgaggccg ccgaggcccc aaagcttcca ctcgatcaat ggtttcctga
      781 ggatgccgac gctgatccta caggtccaaa gagctggtgg ggaagagcta gcgtagacga
      841 taagagagtg tttgtgggtc tatttgttga caagattgta gttactaagt ccactacggg
      901 tcgaggacag ggcactccca tcgaaaagag agccagtatt acctgggcaa agcctccaac
      961 cgatgatgat gaagacgatg cacaagatgg taccgaggat gtggcagcac caaaaaaaaa
     1021 gaggaaggtg taagctttga gacctcatca cacctgaatt ccagcacact ggcggccgtt
     1081 actagtggat ccgagctcgg taccaagctt gatgcatagc ttgagtattc taacgcgtca
     1141 cctaaatagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct
     1201 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg
     1261 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     1321 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     1381 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     1441 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     1501 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     1561 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     1621 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     1681 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     1741 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     1801 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     1861 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     1921 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     1981 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     2041 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     2101 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     2161 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     2221 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     2281 ttttagcacg tgtcagtcct gctcctcggc cacgaagtgc acgcagttgc cggccgggtc
     2341 gcgcagggcg aactcccgcc cccacggctg ctcgccgatc tcggtcatgg ccggcccgga
     2401 ggcgtcccgg aagttcgtgg acacgacctc cgaccactcg gcgtacagct cgtccaggcc
     2461 gcgcacccac acccaggcca gggtgttgtc cggcaccacc tggtcctgga ccgcgctgat
     2521 gaacagggtc acgtcgtccc ggaccacacc ggcgaagtcg tcctccacga agtcccggga
     2581 gaacccgagc cggtcggtcc agaactcgac cgctccggcg acgtcgcgcg cggtgagcac
     2641 cggaacggca ctggtcaact tggccatggt ggccctcctc acgtgctatt attgaagcat
     2701 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     2761 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gatgcggtgt gaaataccgc
     2821 acagatgcgt aaggagaaaa taccgcatca ggaaattgta agcgttaata attcagaaga
     2881 actcgtcaag aaggcgatag aaggcgatgc gctgcgaatc gggagcggcg ataccgtaaa
     2941 gcacgaggaa gcggtcagcc cattcgccgc caagctcttc agcaatatca cgggtagcca
     3001 acgctatgtc ctgatagcgg tccgccacac ccagccggcc acagtcgatg aatccagaaa
     3061 agcggccatt ttccaccatg atattcggca agcaggcatc gccatgggtc acgacgagat
     3121 cctcgccgtc gggcatgctc gccttgagcc tggcgaacag ttcggctggc gcgagcccct
     3181 gatgctcttc gtccagatca tcctgatcga caagaccggc ttccatccga gtacgtgctc
     3241 gctcgatgcg atgtttcgct tggtggtcga atgggcaggt agccggatca agcgtatgca
     3301 gccgccgcat tgcatcagcc atgatggata ctttctcggc aggagcaagg tgagatgaca
     3361 ggagatcctg ccccggcact tcgcccaata gcagccagtc ccttcccgct tcagtgacaa
     3421 cgtcgagcac agctgcgcaa ggaacgcccg tcgtggccag ccacgatagc cgcgctgcct
     3481 cgtcttgcag ttcattcagg gcaccggaca ggtcggtctt gacaaaaaga accgggcgcc
     3541 cctgcgctga cagccggaac acggcggcat cagagcagcc gattgtctgt tgtgcccagt
     3601 catagccgaa tagcctctcc acccaagcgg ccggagaacc tgcgtgcaat ccatcttgtt
     3661 caatcatgcg aaacgatcct catcctgtct cttgatcaga gcttgatccc ctgcgccatc
     3721 agatccttgg cggcaagaaa gccatccagt ttactttgca gggcttccca accttaccag
     3781 agggcgcccc agctggcaat tccggttcgc ttgctgtcca taaaaccgcc cagtctagct
     3841 atcgccatgt aagcccactg caagctacct gctttctctt tgcgcttgcg ttttcccttg
     3901 tccagatagc ccagtagctg acattcatcc ggggtcagca ccgtttctgc ggactggctt
     3961 tctacgtgaa aaggatctag gtgaagatcc tttttgataa tctcatgcct gacatttata
     4021 ttccccagaa catcaggtta atggcgtttt tgatgtcatt ttcgcggtgg ctgagatcag
     4081 ccacttcttc cccgataacg gagaccggca cactggccat atcggtggtc atcatgcgcc
     4141 agctttcatc cccgatatgc accaccgggt aaagttcacg ggagacttta tctgacagca
     4201 gacgtgcact ggccaggggg atcaccatcc gtcgccccgg cgtgtcaata atatcactct
     4261 gtacatccac aaacagacga taacggctct ctcttttata ggtgtaaacc ttaaactgcc
     4321 gtacgtatag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac
     4381 gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt
     4441 cccagtcacg acgttgtaaa acgacggcca gtgaattgta atacgactca ctatagggcg
     4501 aattgggccc tctagatgca tgctcgagcg gccgccagtg tgatggatat ctgcagaatt
     4561 cagg
//
