LOCUS       sequence-485723-        4745 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bxb1-GA donor plasmid for cloning of payloads.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4745)
  AUTHORS   Blanch-Asensio A, Ploessl DS, Wang NB, Mummery CL, Galloway KE, 
            Davis RP
  TITLE     STRAIGHT-IN Dual: a platform for dual, single-copy integrations of 
            DNA payloads and gene circuits into human induced pluripotent stem 
            cells.
  JOURNAL   bioRxiv [Preprint]. 2024 Oct 17:2024.10.17.616637. doi: 
            10.1101/2024.10.17.616637.
  PUBMED    39464154
REFERENCE   2  (bases 1 to 4745)
  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: "bioRxiv 
            [Preprint]. 2024 Oct 17:2024.10.17.616637. doi: 
            10.1101/2024.10.17.616637."
COMMENT     Sequence Label: Bxb1-GA-Donor_UbC
FEATURES             Location/Qualifiers
     source          1..4745
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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)."
     primer_bind     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        373..1584
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     1421..1440
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     protein_bind    1619..1656
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attB site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     protein_bind    1671..1718
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     protein_bind    1803..1836
                     /label=FRT3
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision with other FRT3 sites
                     (Bosch et al., 2015)"
     primer_bind     complement(1844..1860)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1874..1890)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1883..1905)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2103..2558
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2190..2209)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2400..2421
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2575..2594)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2694..2716
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2754..2772)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2840..2944
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2945..3013,3014..3805)
                     /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     complement(3163..3182)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3976..4564
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4465..4484
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4718..4735
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctta
      241 attccctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
      301 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
      361 ccctccagat ctggcctccg cgccgggttt tggcgcctcc cgcgggcgcc cccctcctca
      421 cggcgagcgc tgccacgtca gacgaagggc gcagcgagcg tcctgatcct tccgcccgga
      481 cgctcaggac agcggcccgc tgctcataag actcggcctt agaaccccag tatcagcaga
      541 aggacatttt aggacgggac ttgggtgact ctagggcact ggttttcttt ccagagagcg
      601 gaacaggcga ggaaaagtag tcccttctcg gcgattctgc ggagggatct ccgtggggcg
      661 gtgaacgccg atgattatat aaggacgcgc cgggtgtggc acagctagtt ccgtcgcagc
      721 cgggatttgg gtcgcggttc ttgtttgtgg atcgctgtga tcgtcacttg gtgagtagcg
      781 ggctgctggg ctggccgggg ctttcgtggc cgccgggccg ctcggtggga cggaagcgtg
      841 tggagagacc gccaagggct gtagtctggg tccgcgagca aggttgccct gaactggggg
      901 ttggggggag cgcagcaaaa tggcggctgt tcccgagtct tgaatggaag acgcttgtga
      961 ggcgggctgt gaggtcgttg aaacaaggtg gggggcatgg tgggcggcaa gaacccaagg
     1021 tcttgaggcc ttcgctaatg cgggaaagct cttattcggg tgagatgggc tggggcacca
     1081 tctggggacc ctgacgtgaa gtttgtcact gactggagaa ctcggtttgt cgtctgttgc
     1141 gggggcggca gttatggcgg tgccgttggg cagtgcaccc gtacctttgg gagcgcgcgc
     1201 cctcgtcgtg tcgtgacgtc acccgttctg ttggcttata atgcagggtg gggccacctg
     1261 ccggtaggtg tgcggtaggc ttttctccgt cgcaggacgc agggttcggg cctagggtag
     1321 gctctcctga atcgacaggc gccggacctc tggtgagggg agggataagt gaggcgtcag
     1381 tttctttggt cggttttatg tacctatctt cttaagtagc tgaagctccg gttttgaact
     1441 atgcgctcgg ggttggcgag tgtgttttgt gaagtttttt aggcaccttt tgaaatgtaa
     1501 tcatttgggt caatatgtaa ttttcagtgt tagactagta aattgtccgc taaattctgg
     1561 ccgtttttgg cttttttgtt agacgaagct tgggcccggg aattaattca ccatggccgg
     1621 cttgtcgacg acggcggact ccgtcgtcag gatcatccgg atccggatcc gaagttccta
     1681 ttccgaagtt cctattctct agaaagtata ggaacttcat gatattcggc aagcaggcat
     1741 cgactagtta attaagctag cgctgcaaga actcttcctc acggatccga agttcctatt
     1801 ccgaagttcc tattcttcaa atagtatagg aacttcagcg ctcactggcc gtcgttttac
     1861 aaagggcgaa ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta
     1921 cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg
     1981 cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg cgcctgatgc
     2041 ggtattttct ccttacgcat ctgtgcggta tttcacaccg catacgtcaa agcaaccata
     2101 gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     2161 cgctacactt gccagcgcct tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     2221 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     2281 tagtgcttta cggcacctcg accccaaaaa acttgatttg ggtgatggtt cacgtagtgg
     2341 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     2401 tggactcttg ttccaaactg gaacaacact caactctatc tcgggctatt cttttgattt
     2461 ataagggatt ttgccgattt cggtctattg gttaaaaaat gagctgattt aacaaaaatt
     2521 taacgcgaat tttaacaaaa tattaacgtt tacaatttta tggtgcactc tcagtacaat
     2581 ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc
     2641 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag
     2701 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt
     2761 gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg
     2821 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     2881 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     2941 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     3001 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     3061 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     3121 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     3181 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     3241 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     3301 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     3361 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     3421 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     3481 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     3541 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     3601 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     3661 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     3721 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     3781 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     3841 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     3901 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     3961 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     4021 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     4081 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     4141 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     4201 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     4261 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     4321 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     4381 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     4441 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     4501 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     4561 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     4621 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     4681 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
     4741 ggaag
//
