LOCUS       sequence-364562-        5801 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Bxb1 integrase in mammalian cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5801)
  AUTHORS   Zhu F, Gamboa M, Farruggio AP, Hippenmeyer S, Tasic B, Schule B, 
            Chen-Tsai Y, Calos MP
  TITLE     DICE, an efficient system for iterative genomic editing in human 
            pluripotent stem cells.
  JOURNAL   Nucleic Acids Res. 2013 Dec 4.
  PUBMED    24304893
REFERENCE   2  (bases 1 to 5801)
  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: "Nucleic 
            Acids Res. 2013 Dec 4."
COMMENT     Sequence Label: pCMV-Bx
FEATURES             Location/Qualifiers
     source          1..5801
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(66..83)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(237..825)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(317..336)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    complement(903..936)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(1038..1829,1830..1898))
                     /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     1661..1680
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(2185..2639)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2322..2343)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2533..2552
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    2765..2899
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2789..2808)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2843..2862
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(3311..4813)
                     /codon_start=1
                     /transl_table=11
                     /gene="Bxb1"
                     /product="Bxb1 integrase"
                     /label=Bxb1 integrase
                     /note="Bxb1 large serine integrase from Mycobacterium 
                     bacteriophage Bxb1 (Ghosh et al. 2003)."
                     /protein_id="AFF61396.1"
                     /translation="MRALVVIRLSRVTDATTSPERQLESCQQLCAQRGWDVVGVAEDLD
                     VSGAVDPFDRKRRPNLARWLAFEEQPFDVIVAYRVDRLTRSIRHLQQLVHWAEDHKKLV
                     VSATEAHFDTTTPFAAVVIALMGTVAQMELEAIKERNRSAAHFNIRAGKYRGSLPPWGY
                     LPTRVDGEWRLVPDPVQRERILEVYHRVVDNHEPLHLVAHDLNRRGVLSPKDYFAQLQG
                     REPQGREWSATALKRSMISEAMLGYATLNGKTVRDDDGAPLVRAEPILTREQLEALRAE
                     LVKTSRAKPAVSTPSLLLRVLFCAVCGEPAYKFAGGGRKHPRYRCRSMGFPKHCGNGTV
                     AMAEWDAFCEEQVLDLLGDAERLEKVWVAGSDSAVELAEVNAELVDLTSLIGSPAYRAG
                     SPQREALDARIAALAARQEELEGLEARPSGWEWRETGQRFGDWWREQDTAAKNTWLRSM
                     NVRLTFDVRGGLTRTIDFGDLQEYEQHLRLGSVVERLHTGMS"
     promoter        complement(4851..4869)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(4852..4869)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(4964..4988)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        complement(4985..5188)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(5014..5034)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        5189..5492
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     complement(5511..5533)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5522..5538
                     /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(5546..5552,5553..5570,5571..5576))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     primer_bind     complement(5603..5621)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5689..5793
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
       61 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
      121 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
      181 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
      241 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
      301 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
      361 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
      421 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
      481 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
      541 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
      601 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
      661 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
      721 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
      781 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
      841 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
      901 ccataacttc gtatagcata cattatacga agttatggca tgagattatc aaaaaggatc
      961 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     1021 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     1081 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     1141 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     1201 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     1261 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     1321 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     1381 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     1441 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     1501 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     1561 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     1621 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     1681 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     1741 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     1801 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     1861 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     1921 tgaagcattt atcagggtta ttgtctcatg ccaggggtgg gcacacatat ttgataccag
     1981 cgatccctac acagcacata attcaatgcg acttccctct atcgcacatc ttagaccttt
     2041 attctccctc cagcacacat cgaagctgcc gagcaagccg ttctcaccag tccaagacct
     2101 ggcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     2161 acatttcccc gaaaagtgcc acctaaattg taagcgttaa tattttgtta aaattcgcgt
     2221 taaatttttg ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt
     2281 ataaatcaaa agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc
     2341 cactattaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
     2401 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     2461 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     2521 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     2581 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     2641 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
     2701 attacgccag ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
     2761 gatcgatcca gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca
     2821 gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat
     2881 aagctgcaat aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg
     2941 ggaggtgtgg gaggtttttt aaagcaagta aaacctctac aaatgtggta tggctgatta
     3001 tgatcatgaa cagactgtga ggactgaggg gcctgaaatg agccttggga ctgtgaatct
     3061 aaaatacaca aacaattaga atcactagct cctgtgtata atattttcat aaatcatact
     3121 cagtaagcaa aactctcaag cagcaagcat atgcagctag tttaacacat tatacactta
     3181 aaaattttat atttacctta gagctttaaa tctctgtagg tagtttgtcc aattatgtca
     3241 caccacagaa gtaaggttcc ttcacaaaga tcccaagcta gtgagctcgt cgacccggga
     3301 attcgccctt ctacgacatc ccggtgtgta gccgttcgac cacgctgccg agcctgagat
     3361 gctgctcgta ctcctgcaga tccccgaagt cgatcgtgcg agtcagcccg ccgcggacgt
     3421 cgaacgtcag ccgaacgttc atcgaccgaa gccaggtgtt ctttgccgcg gtgtcctgct
     3481 cccgccacca gtccccgaac cgctgcccgg tctcgcgcca ctcccagcca gacgggcgag
     3541 cctctagacc ctccagctcc tcttgccgcg cggccagcgc cgcaatacgg gcatccagtg
     3601 cttctcgctg cggagagccg gcccggtagg ccggggagcc gatcagcgac gtcaggtcca
     3661 ccagctccgc gttcacctcc gcgagttcga ccgcggagtc cgagccggct acccagactt
     3721 tctccagacg ctccgcgtcc ccgagcagat ccagcacctg ctcctcgcag aacgcgtccc
     3781 actcggccat cgccaccgtg ccgttcccgc agtgcttcgg gaaccccatc gagcggcagc
     3841 ggtagcgcgg gtgcttacgt cctcccccgg cgaacttgta cgcgggctcc ccgcacaccg
     3901 cgcagaacaa cacccgcagc agcagcgacg gggtagacac cgcgggcttc gcccgggagg
     3961 tcttcacgag ctcggcgcgc agcgcctcca gctgctcacg ggtcaggatc ggctcagccc
     4021 gcaccagcgg ggctccgtcg tcgtctcgga cggtcttacc gttcagagtc gcgtacccga
     4081 gcatcgcctc ggagatcatc gatcgcttca gcgcggtagc cgaccactcc cggccctgcg
     4141 gctcgcggcc ttgcagctgc gcgaagtagt ccttcggcga caggacacca cgccggttca
     4201 ggtcgtgggc caccaggtgc agcggctcgt ggttgtcgac gacgcggtga tacacctcga
     4261 ggatgcgctc tcgctgcaca gggtccggca ccagccgcca ctccccgtcc acgcgcgtag
     4321 gcaggtatcc ccacggcggc agggatcctc ggtatttccc ggcgcggata ttgaaatgcg
     4381 cagccgaacg gttccgctct ttgatcgctt ctaattccat ctgcgccacc gttcccataa
     4441 gcgcgatgac gaccgccgca aacggcgtcg tcgtatcgaa gtgcgcttcg gtcgcggaga
     4501 cgaccagctt cttgtggtcc tcggcccagt ggaccagctg ctgcagatgc cggatcgatc
     4561 gggtcaaccg gtctacccgg tacgccacga tcacgtcgaa cggttgctcc tcgaacgcta
     4621 gccaccgggc caggttcggt ctgcgcttcc ggtcgaacgg atcgaccgcc ccggagacgt
     4681 ccagatcctc cgctaccccg acgacgtccc agccgcgctg ggcgcagagc tgctggcaag
     4741 actccagctg acgctccggt gaagtcgtag catcggtgac gcgggacagg cggatgacta
     4801 ccagggctct cataagggcg aattccggac cggtacctgc aggcgtacct tctatagtgt
     4861 cacctaaata gctttttgca aaagcctagg ctagagtccg gaggctggat cggtcccggt
     4921 gtcttctatg gaggtcaaaa cagcgtggat ggcgcctcca ggcgatctga cggttcacta
     4981 aacgagctct gcttatatag acctcccacc gtacacgcct accgcccatt tgcgtcaatg
     5041 gggcggagtt gttacgacat tttggaaagt cccgttgatt ttggtgccaa aacaaactcc
     5101 cattgacgtc aatggggtgg agacttggaa atccccgtga gtcaaaccgc tatccacgcc
     5161 cattgatgta ctgccaaaac cgcatcacca tggtaatagc gatgactaat acgtagatgt
     5221 actgccaagt aggaaagtcc cataaggtca tgtactgggc ataatgccag gcgggccatt
     5281 taccgtcatt gacgtcaata gggggcgtac ttggcatatg atacacttga tgtactgcca
     5341 agtgggcagt ttaccgtaaa tactccaccc attgacgtca atggaaagtc cctattggcg
     5401 ttactatggg aacatacgtc attattgacg tcaatgggcg ggggtcgttg ggcggtcagc
     5461 caggcgggcc atttaccgta agttatgtaa cgacctgcac gatgctgttt cctgtgtgaa
     5521 attgttatcc gctcacaatt ccacacatta tacgagccgg aagcataaag tgtaaagcct
     5581 ggggtgccta atgagtgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg
     5641 ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
     5701 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     5761 taaatgcttc aataatattg aaaaagcgcg aattgcaagc t
//
