LOCUS       sequence-262666-        7186 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BxBI Destination vector with H2B::GFP expressed from CAG promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7186)
  AUTHORS   Fonseca JP, Bonny AR, Kumar GR, Ng AH, Town J, Wu QC, Aslankoohi E, 
            Chen SY, Dods G, Harrigan P, Osimiri LC, Kistler AL, El-Samad H
  TITLE     A Toolkit for Rapid Modular Construction of Biological Circuits in 
            Mammalian Cells
  JOURNAL   ACS Synth Biol. 2019 Nov 15;8(11):2593-2606. doi: 
            10.1021/acssynbio.9b00322. Epub 2019 Nov 5.
  PUBMED    31686495
REFERENCE   2  (bases 1 to 7186)
  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"; doi: 
            "10.1021/acssynbio.9b00322"; journalName: "ACS Synth Biol"; date: 
            "2019-11-15- 15"; volume: "8"; issue: "11"; pages: "2593-2606"
COMMENT     Sequence Label: JPF0446
FEATURES             Location/Qualifiers
     source          1..7186
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    637..674
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attB site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     CDS             683..1381
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     CDS             1382..1402
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1433..1486
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             1493..1891
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    1909..2116
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     CDS             complement(2171..2986)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2171..2986)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2894..2913
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2978..2997
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3211..3799
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3700..3719
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    3911..3944
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     enhancer        4174..4477
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4479..4754
                     /label=chicken beta-actin promoter
     intron          4755..5771
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     5695..5717
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     5779..5798
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             5832..6209
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     CDS             join(6216..6218,6219..6932)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(6258..6279)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     6866..6887
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    6954..7178
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 gttacagaga tgttacgaac cactagtgca ctgcagatcg aattcgagtt ggcgcgcctg
       61 tcattctaaa tctctctttc agcctaaagc tttttccccg tatcccccca ggtgtctgca
      121 ggctcaaaga gcagcgagaa gcgttcagag gaaagcgatc ccgtgccacc ttccccgtgc
      181 ccgggctgtc cccgcacgct gccggctcgg ggatgcgggg ggagcgccgg accggagcgg
      241 agccccgggc ggctcgctgc tgccccctag cgggggaggg acgtaattac atccctgggg
      301 gctttgggag ggggctgtcc ccgtgagctc ccaggcgcgc ctgtcattct aaatctctct
      361 ttcagcctaa agctttttcc ccgtatcccc ccaggtgtct gcaggctcaa agagcagcga
      421 gaagcgttca gaggaaagcg atcccgtgcc accttccccg tgcccgggct gtccccgcac
      481 gctgccggct cggggatgcg gggggagcgc cggaccggag cggagccccg ggcggctcgc
      541 tgctgccccc tagcggggga gggacgtaat tacatccctg ggggctttgg gggggggctg
      601 tccccgtgag ctcaataaaa gagcccacta caggccggct tgtcgacgac ggcggtctcc
      661 gtcgtcagga tcatccggga gtatgagcga gctgattaag gagaacatgc acatgaagct
      721 gtacatggag ggcaccgtgg acaaccatca cttcaagtgc acatccgagg gcgaaggcaa
      781 gccctacgag ggcacccaga ccatgagaat caaggtggtc gagggcggcc ctctcccctt
      841 cgccttcgac atcctggcta ctagcttcct ctacggcagc aagaccttca tcaaccacac
      901 ccagggcatc cccgacttct tcaagcagtc cttccctgag ggcttcacat gggagagagt
      961 caccacatac gaagacgggg gcgtgctgac cgctacccag gacaccagcc tccaggacgg
     1021 ctgcctcatc tacaacgtca agatcagagg ggtgaacttc acatccaacg gccctgtgat
     1081 gcagaagaaa acactcggct gggaggcctt caccgaaacg ctgtaccccg ctgacggcgg
     1141 cctggaaggc agaaacgaca tggccctgaa gctcgtgggc gggagccatc tgatcgcaaa
     1201 catcaagacc acatatagat ccaagaaacc cgctaagaac ctcaagatgc ctggcgtcta
     1261 ctatgtggac tacagactgg aaagaatcaa ggaggccaac aacgaaacct acgtcgagca
     1321 gcacgaggtg gcagtggcca gatactgcga cctccctagc aaactggggc acaagcttaa
     1381 tcccaagaag aagaggaagg tggaggacgc ccgtacgagt actggaagcg gagagggcag
     1441 aggaagtctg ctaacatgcg gtgacgtcga ggagaatcct ggacctggat ccatggccaa
     1501 gcctttgtct caagaagaat ccaccctcat tgaaagagca acggctacaa tcaacagcat
     1561 ccccatctct gaagactaca gcgtcgccag cgcagctctc tctagcgacg gccgcatctt
     1621 cactggtgtc aatgtatatc attttactgg gggaccttgt gcagaactcg tggtgctggg
     1681 cactgctgct gctgcggcag ctggcaacct gacttgtatc gtcgcgatcg gaaatgagaa
     1741 caggggcatc ttgagcccct gcggacggtg ccgacaggtg cttctcgatc tgcatcctgg
     1801 gatcaaagcc atagtgaagg acagtgatgg acagccgacg gcagttggga ttcgtgaatt
     1861 gctgccctct ggttatgtgt gggagggcta actcgagcga cgcctcgact gtgccttcta
     1921 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca
     1981 ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc
     2041 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata
     2101 gcaggcatgc tggggatgcg gtgggctcta tccgagcggc cgcgtgttac aaccaattaa
     2161 ccaattctga ttagaaaaac tcatcgagca tcaaatgaaa ctgcaattta ttcatatcag
     2221 gattatcaat accatatttt tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga
     2281 ggcagttcca taggatggca agatcctggt atcggtctgc gattccgact cgtccaacat
     2341 caatacaacc tattaatttc ccctcgtcaa aaataaggtt atcaagtgag aaatcaccat
     2401 gagtgacgac tgaatccggt gagaatggca aaagcttatg catttctttc cagacttgtt
     2461 caacaggcca gccattacgc tcgtcatcaa aatcactcgc atcaaccaaa ccgttattca
     2521 ttcgtgattg cgcctgagcg aggcgaaata cgcgatcgct gttaaaagga caattacaaa
     2581 caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc atcaacaata ttttcacctg
     2641 aatcaggata ttcttctaat acctggaatg ctgttttccc ggggatcgca gtggtgagta
     2701 accatgcatc atcaggagta cggataaaat gcttgatggt cggaagaggc ataaattccg
     2761 tcagccagtt tagtctgacc atctcatctg taacatcatt ggcaacgcta cctttgccat
     2821 gtttcagaaa caactctggc gcatcgggct tcccatacaa tcgatagatt gtcgcacctg
     2881 attgcccgac attatcgcga gcccatttat acccatataa atcagcatcc atgttggaat
     2941 ttaatcgcgg cctggagcaa gacgtttccc gttgaatatg gctcataaca ccccttgtat
     3001 tactgtttat gtaagcagac agttttattg ttcatgatga tatattttta tcttgtgcaa
     3061 tgtaacatca gagattttga gacacaacgt ggctttgttg aataaatcga acttttgctg
     3121 agttgaagga tcagtcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     3181 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     3241 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     3301 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     3361 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     3421 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     3481 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     3541 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     3601 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     3661 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     3721 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     3781 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     3841 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtgc
     3901 ggccgccaat ataacttcgt ataatgtatg ctatacgaag ttatccctga attcgcatct
     3961 agactgaact ggccgataat tgcagacgag gagcatcgcc cttccccggc cctcaggtaa
     4021 gaggaccaaa taccgtagcc gtttccaatt tcagtccttt agcgccacct ggtgctaact
     4081 actctatcac gcttttatcc aataactacc tttgtaaatt tcctttcaaa agttctggcc
     4141 gggcgcggtg gcaacgccat atatggagtt ccgcgttaca taacttacgg taaatggccc
     4201 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
     4261 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
     4321 ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga
     4381 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg
     4441 gcagtacatc tacgtattag tcatcgctat taccatggtc gaggtgagcc ccacgttctg
     4501 cttcactctc cccatctccc ccccctcccc acccccaatt ttgtatttat ttatttttta
     4561 attattttgt gcagcgatgg gggcgggggg gggggggggg cgcgccaggc ggggcggggc
     4621 ggggcgaggg gcggggcggg gcgaggcgga gaggtgcggc ggcagccaat cagagcggcg
     4681 cgctccgaaa gtttcctttt atggcgaggc ggcggcggcg gcggccctat aaaaagcgaa
     4741 gcgcgcggcg ggcgggagtc gctgcgcgct gccttcgccc cgtgccccgc tccgccgccg
     4801 cctcgcgccg cccgccccgg ctctgactga ccgcgttact cccacaggtg agcgggcggg
     4861 acggcccttc tcctccgggc tgtaattagc gcttggttta atgacggctt gtttcttttc
     4921 tgtggctgcg tgaaagcctt gaggggctcc gggagggccc tttgtgcggg gggagcggct
     4981 cggggggtgc gtgcgtgtgt gtgtgcgtgg ggagcgccgc gtgcggctcc gcgctgcccg
     5041 gcggctgtga gcgctgcggg cgcggcgcgg ggctttgtgc gctccgcagt gtgcgcgagg
     5101 ggagcgcggc cgggggcggt gccccgcggt gcgggggggg ctgcgagggg aacaaaggct
     5161 gcgtgcgggg tgtgtgcgtg ggggggtgag cagggggtgt gggcgcgtcg gtcgggctgc
     5221 aaccccccct gcacccccct ccccgagttg ctgagcacgg cccggcttcg ggtgcggggc
     5281 tccgtacggg gcgtggcgcg gggctcgccg tgccgggcgg ggggtggcgg caggtggggg
     5341 tgccgggcgg ggcggggccg cctcgggccg gggagggctc gggggagggg cgcggcggcc
     5401 cccggagtgc cggcggctgt cgaggcgcgg cgagccgcag ccattgcctt ttatggtaat
     5461 cgtgcgagag ggcgcaggga cttcctttgt cccaaatctg tgcggagccg aaatctggga
     5521 ggcgccgccg caccccctct agcgggcgcg gggcgaagcg gtgcggcgcc ggcaggaagg
     5581 aaatgggcgg ggagggcctt cgtgcgtcgc cgcgccgccg tccccttctc cctctccagc
     5641 ctcggggctg tccgcggggg gacggctgcc ttcggggggg acggggcagg gcggggttcg
     5701 gcttctggcg tgtgaccggc ggctctagag cctctgctaa ccatgttcat gccttcttct
     5761 ttttcctaca gctcctgggc aacgtgctgg ttattgtgct gtctcatcat tttggcaaag
     5821 aattcgccac tatgccagag ccagcgaagt ctgctcccgc cccgaaaaag ggctccaaga
     5881 aggcggtgac taaggcgcag aagaaaggcg gcaagaagcg caagcgcagc cgcaaggaga
     5941 gctattccat ctatgtgtac aaggttctga agcaggtcca ccctgacacc ggcatttcgt
     6001 ccaaggccat gggcatcatg aattcgtttg tgaacgacat tttcgagcgc atcgcaggtg
     6061 aggcttcccg cctggcgcat tacaacaagc gctcgaccat cacctccagg gagatccaga
     6121 cggccgtgcg cctgctgctg cctggggagt tggccaagca cgccgtgtcc gagggtacta
     6181 aggccatcac caagtacacc agcgctaagg gatccgtgag caagggcgag gagctgttca
     6241 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
     6301 tgcgcggcga gggcgagggc gatgccacca acggcaagct gaccctgaag ttcatctgca
     6361 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc
     6421 agtgcttcag ccgctacccc gaccacatga agcgccacga cttcttcaag tccgccatgc
     6481 ccgaaggcta cgtccaggag cgcaccatca gcttcaagga cgacggcacc tacaagaccc
     6541 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     6601 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaacttc aacagccaca
     6661 acgtctatat caccgccgac aagcagaaga acggcatcaa ggccaacttc aagatccgcc
     6721 acaacgtgga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     6781 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc aagctgagca
     6841 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     6901 tcactcacgg catggacgag ctgtacaagt aactcgagtg gccgacgcct cgactgtgcc
     6961 ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg
     7021 tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag
     7081 gtgtcattct attctggggg gtggggtggt gcaggacagc aagggggagg attgggaaga
     7141 caatagcagg catgctgggg atgcggtggg ctctatgggc tgagca
//
