LOCUS       sequence-426688-        3757 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level 0 Empty Bxb1 target: attP_Bxb1-BpiI-attB_Bxb1 with golden gate
            linker in pBLUNT plasmid..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3757)
  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 3757)
  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-T6 Empty target Bxb1
FEATURES             Location/Qualifiers
     source          1..3757
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    24..71
                     /label=Bxb1 attP
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attP site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     protein_bind    137..186
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="attB site for the Mycopbacteriophage Bxb1 integrase 
                     (Xu et al., 2013)"
     primer_bind     complement(318..334)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(318..334)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(331..353)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    342..358
                     /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(366..372,373..390,391..396))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    411..432
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(549..566)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(720..1308)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(800..819)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1446..1820)
                     /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      1817..1826
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(2026..2820)
                     /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(2026..2820)
                     /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(2137..2156)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     2747..2766
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     CDS             complement(3169..3471)
                     /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(3271..3290)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     primer_bind     3594..3616
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3608..3625
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3609..3625
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3632..3651
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3632..3650
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 cactctgtgg tctcactatt cgtggtttgt ctggtcaacc accgccgtct cagtggtgta
       61 cggtacaaac ccggagttgt cttcttgtag cactgtaaga tttatccacg aaggtcagca
      121 acttgaagac aatactgccc ggatgatcct gacgacggag acggccgtcg tcgacaagcc
      181 ggccgaacct tgagacctcc gatcccctga attccagcac actggcggcc gttactagtg
      241 gatccgagct cggtaccaag cttgatgcat agcttgagta ttctaacgcg tcacctaaat
      301 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
      361 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
      421 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
      481 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
      541 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
      601 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
      661 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
      721 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
      781 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
      841 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
      901 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
      961 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     1021 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     1081 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     1141 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     1201 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     1261 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     1321 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     1381 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttagc
     1441 acgtgtcagt cctgctcctc ggccacgaag tgcacgcagt tgccggccgg gtcgcgcagg
     1501 gcgaactccc gcccccacgg ctgctcgccg atctcggtca tggccggccc ggaggcgtcc
     1561 cggaagttcg tggacacgac ctccgaccac tcggcgtaca gctcgtccag gccgcgcacc
     1621 cacacccagg ccagggtgtt gtccggcacc acctggtcct ggaccgcgct gatgaacagg
     1681 gtcacgtcgt cccggaccac accggcgaag tcgtcctcca cgaagtcccg ggagaacccg
     1741 agccggtcgg tccagaactc gaccgctccg gcgacgtcgc gcgcggtgag caccggaacg
     1801 gcactggtca acttggccat ggtggccctc ctcacgtgct attattgaag catttatcag
     1861 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     1921 gttccgcgca catttccccg aaaagtgcca cctgatgcgg tgtgaaatac cgcacagatg
     1981 cgtaaggaga aaataccgca tcaggaaatt gtaagcgtta ataattcaga agaactcgtc
     2041 aagaaggcga tagaaggcga tgcgctgcga atcgggagcg gcgataccgt aaagcacgag
     2101 gaagcggtca gcccattcgc cgccaagctc ttcagcaata tcacgggtag ccaacgctat
     2161 gtcctgatag cggtccgcca cacccagccg gccacagtcg atgaatccag aaaagcggcc
     2221 attttccacc atgatattcg gcaagcaggc atcgccatgg gtcacgacga gatcctcgcc
     2281 gtcgggcatg ctcgccttga gcctggcgaa cagttcggct ggcgcgagcc cctgatgctc
     2341 ttcgtccaga tcatcctgat cgacaagacc ggcttccatc cgagtacgtg ctcgctcgat
     2401 gcgatgtttc gcttggtggt cgaatgggca ggtagccgga tcaagcgtat gcagccgccg
     2461 cattgcatca gccatgatgg atactttctc ggcaggagca aggtgagatg acaggagatc
     2521 ctgccccggc acttcgccca atagcagcca gtcccttccc gcttcagtga caacgtcgag
     2581 cacagctgcg caaggaacgc ccgtcgtggc cagccacgat agccgcgctg cctcgtcttg
     2641 cagttcattc agggcaccgg acaggtcggt cttgacaaaa agaaccgggc gcccctgcgc
     2701 tgacagccgg aacacggcgg catcagagca gccgattgtc tgttgtgccc agtcatagcc
     2761 gaatagcctc tccacccaag cggccggaga acctgcgtgc aatccatctt gttcaatcat
     2821 gcgaaacgat cctcatcctg tctcttgatc agagcttgat cccctgcgcc atcagatcct
     2881 tggcggcaag aaagccatcc agtttacttt gcagggcttc ccaaccttac cagagggcgc
     2941 cccagctggc aattccggtt cgcttgctgt ccataaaacc gcccagtcta gctatcgcca
     3001 tgtaagccca ctgcaagcta cctgctttct ctttgcgctt gcgttttccc ttgtccagat
     3061 agcccagtag ctgacattca tccggggtca gcaccgtttc tgcggactgg ctttctacgt
     3121 gaaaaggatc taggtgaaga tcctttttga taatctcatg cctgacattt atattcccca
     3181 gaacatcagg ttaatggcgt ttttgatgtc attttcgcgg tggctgagat cagccacttc
     3241 ttccccgata acggagaccg gcacactggc catatcggtg gtcatcatgc gccagctttc
     3301 atccccgata tgcaccaccg ggtaaagttc acgggagact ttatctgaca gcagacgtgc
     3361 actggccagg gggatcacca tccgtcgccc cggcgtgtca ataatatcac tctgtacatc
     3421 cacaaacaga cgataacggc tctctctttt ataggtgtaa accttaaact gccgtacgta
     3481 taggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct
     3541 ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc
     3601 acgacgttgt aaaacgacgg ccagtgaatt gtaatacgac tcactatagg gcgaattggg
     3661 ccctctagat gcatgctcga gcggccgcca gtgtgatgga tatctgcaga attcaggcga
     3721 gcggccgcca gtgtgatgga tatctgcaga attcagg
//
