LOCUS       sequence-461267-        4360 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  bigMamAct is evolution of biGBac for mammalian cells. psBIG plasmids
            are recipient of multi-assembly step by Gibson cloning .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4360)
  AUTHORS   Pelosse M, Marcia M
  TITLE     biGMamAct: efficient CRISPR/Cas9-mediated docking of large 
            functional DNA cargoes at the ACTB locus
  JOURNAL   Synthetic Biology 2025.02.13
REFERENCE   2  (bases 1 to 4360)
  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: "Synthetic 
            Biology 2025.02.13"
COMMENT     Sequence Label: psBIG1d
FEATURES             Location/Qualifiers
     source          1..4360
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             442..1233
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      1335..1923
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1824..1843
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2077..2094
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  2228..2452
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(2519..3052)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3241..3246,3247..3263,3264..3269))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     protein_bind    complement(3665..3698)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(3744..3763)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3788..3836
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     mobile_element  4113..4278
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
ORIGIN
        1 ttctctgtca cagaatgaaa atttttctgt catctcttcg ttattaatgt ttgtaattga
       61 ctgaatatca acgcttattt gcagcctgaa ctgggaggca gaataaatga tcatatcgtc
      121 aattattacc tccacgggga gagcctgagc aaactggcct caggcatttg agaagcacac
      181 ggtcacactg cttccggtag tcaataaacc ggtaagtagc gtatgcgctc acgcaactgg
      241 tccagaacct tgaccgaacg cagcggtggt aacggcgcag tggcggtttt catggcttgt
      301 tatgactgtt tttttggggt acagtctatg cctcgggcat ccaagcagca agcgcgttac
      361 gccgtgggtc gatgtttgat gttatggagc agcaacgatg ttacgcagca gggcagtcgc
      421 cctaaaacaa agttaaacat catgagggaa gcggtgatcg ccgaagtatc gactcaacta
      481 tcagaggtag ttggcgtcat cgagcgccat ctcgaaccga cgttgctggc cgtacatttg
      541 tacggctccg cagtggatgg cggcctgaag ccacacagtg atattgattt gctggttacg
      601 gtgaccgtaa ggcttgatga aacaacgcgg cgagctttga tcaacgacct tttggaaact
      661 tcggcttccc ctggagagag cgagattctc cgcgctgtag aagtcaccat tgttgtgcac
      721 gacgacatca ttccgtggcg ttatccagct aagcgcgaac tgcaatttgg agaatggcag
      781 cgcaatgaca ttcttgcagg tatcttcgag ccagccacga tcgacattga tctggctatc
      841 ttgctgacaa aagcaagaga acatagcgtt gccttggtag gtccagcggc ggaggaactc
      901 tttgatccgg ttcctgaaca ggatctattt gaggcgctaa atgaaacctt aacgctatgg
      961 aactcgccgc ccgactgggc tggcgatgag cgaaatgtag tgcttacgtt gtcccgcatt
     1021 tggtacagcg cagtaaccgg caaaatcgcg ccgaaggatg tcgctgccga ctgggcaatg
     1081 gagcgcctgc cggcccagta tcagcccgtc atacttgaag ctagacaggc ttatcttgga
     1141 caagaagaag atcgcttggc ctcgcgcgca gatcagttgg aagaatttgt ccactacgtg
     1201 aaaggcgaga tcaccaaggt agtcggcaaa taatgtctaa caattcgttc aagccgacgg
     1261 attggcgaat gggacgcgcc cgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     1321 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     1381 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     1441 aaggtaactg gcttcagcag agcgcagata ccaaatacta ttcttctagt gtagccgtag
     1501 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     1561 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     1621 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     1681 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     1741 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     1801 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     1861 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     1921 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     1981 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     2041 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     2101 gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
     2161 gaccagccgc gtaacctggc aaaatcggtt acggttgagt aataaatgga tgccctgcgt
     2221 aagcgggtgt gggcggacaa taaagtctta aactgaacaa aatagatcta aactatgaca
     2281 ataaagtctt aaactagaca gaatagttgt aaactgaaat cagtccagtt atgctgtgaa
     2341 aaagcatact ggacttttgt tatggctaaa gcaaactctt cattttctga agtgcaaatt
     2401 gcccgtcgta ttaaagaggg gcgtggccaa gggcatggta aagactatat tcgcggcgtt
     2461 gtgacaattt accgaacaac tccgcggccg ggaagccgat ctcggcttga acgaattgtt
     2521 aggtggcggt acttgggtcg atatcaaagt gcatcacttc ttcccgtatg cccaactttg
     2581 tatagagagc cactgcggga tcgtcaccgt aatctgcttg cacgtagatc acataagcac
     2641 caagcgcgtt ggcctcatgc ttgaggagat tgatgagcgc ggtggcaatg ccctgcctcc
     2701 ggtgctcgcc ggagactgcg agatcataga tatagatctc actacgcggc tgctcaaact
     2761 tgggcagaac gtaagccgcg agagcgccaa caaccgcttc ttggtcgaag gcagcaagcg
     2821 cgatgaatgt cttactacgg agcaagttcc cgaggtaatc ggagtccggc tgatgttggg
     2881 agtaggtggc tacgtctccg aactcacgac cgaaaagatc aagagcagcc cgcatggatt
     2941 tgacttggtc agggccgagc ctacatgtgc gaatgatgcc catacttgag ccacctaact
     3001 ttgttttagg gcgactgccc tgctgcgtaa catcgttgct gctgcgtaac atcgttgctg
     3061 ctccataaca tcaaacatcg acccacggcg taacgcgctt gctgcttgga tgcccgaggc
     3121 atagactgta caaaaaaaca gtcataacaa gccatgaaaa ccgccactgc gccgttacca
     3181 ccgctgcgtt cggtcaaggt tctggaccag ttgcgtgagc gcatacgcta cttgcattac
     3241 agtttacgaa ccgaacaggc ttatgtcaac tgggtttgtg ccttcatccg tttccacggt
     3301 gtgcgtcacc cggcaacctt gggcagcagc gaagtcgagg catttctgtc ctggctggcg
     3361 aacgagcgca aggtttcggt ctccacgcat cgtcaggcat tggcggcctt gctgttcttc
     3421 tacggcaagg tgctgtgcac ggatctgccc ttgcttcagg agatcggtag acctcggccg
     3481 tcgcggcgct tgccggtggt gctgaccccg gatgaagtgg ttcgcatcct cggttttctg
     3541 gaaggcgagc atcgtttgtt cgcccaggac tctagctata gttctagtgg ttggctacag
     3601 ctttgtttgt actatcaaca ggttgaactg ctgatcaaca gatcctctac gcggccgcgg
     3661 taccataact tcgtatagca tacattatac gaagttatct gggtttaaac ctaatgatgc
     3721 ctgatgtttt tatcgaggct aactgaaaca cggaaggaga caataccgga aggaacccgc
     3781 gctatgacgg caataaaaag acagaataaa acgcacgggt gttgggtcgt ttgttcataa
     3841 acgcggggtt cggtcccagg gctggcactc tgtcaagtga acgctctatg gtctaaagat
     3901 ttaaatctat atctcaatcg gggttacgtc gataccccac cgagacccca ttgggaccaa
     3961 tacgcccgcg tttcttcctt ttccccaccc caacccccaa gttcgggtga aggcccaggg
     4021 ctcgcagcca acgtcggggc ggcaagccct gccatagcca ctacgggtta attaaaaacg
     4081 gttcacatag cttagtttaa acaggagatc cgaaccagat aagtgaaatc tagttccaaa
     4141 ctattttgtc atttttaatt ttcgtattag cttacgacgc tacacccagt tcccatctat
     4201 tttgtcactc ttccctaaat aatccttaaa aactccattt ccacccctcc cagttcccaa
     4261 ctattttgtc cgcccacagc ggggcatttt tcttcctgtt atgtttttaa tcaaacatcc
     4321 tgccaactcc atgtgacaaa ccgtcatctt cggctacttt
//
