LOCUS       sequence-437339-        4984 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for wild type Bxb1 mRNA in vitro transcription template .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4984)
  AUTHORS   Pandey S, Gao XD, Krasnow NA, McElroy A, Tao YA, Duby JE, Steinbeck 
            BJ, McCreary J, Pierce SE, Tolar J, Meissner TB, Chaikof EL, Osborn 
            MJ, Liu DR
  TITLE     Efficient site-specific integration of large genes in mammalian 
            cells via continuously evolved recombinases and prime editing.
  JOURNAL   Nat Biomed Eng. 2024 Jun 10. doi: 10.1038/s41551-024-01227-1.
  PUBMED    38858586
REFERENCE   2  (bases 1 to 4984)
  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 Biomed 
            Eng. 2024 Jun 10. doi: 10.1038/s41551-024-01227-1."
COMMENT     Sequence Label: wtBxb1_IVT
FEATURES             Location/Qualifiers
     source          1..4984
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     polyA_signal    2294..2518
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2589..2605)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2589..2605)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2602..2624)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2613..2629
                     /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(2637..2643,2644..2661,2662..2667))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2682..2703
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2820..2837)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2991..3579)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3071..3090)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3750..4541,4542..4610))
                     /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     4373..4392
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4611..4715)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4794..4813)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc
      601 agatctcgag ctcggtacct aatacgacac actataagga aataagagag aaaagaagag
      661 taagaagaaa tataagagcc accatgcggg ccctggttgt gattagactg tccagagtga
      721 ccgacgccac cacctcacct gagagacagt tggagagctg ccagcagctg tgcgcccagc
      781 gcggctggga cgtggtgggc gtggccgagg acctggacgt gtccggcgcc gtggaccctt
      841 ttgaccggaa gcggagacct aatctggcca gatggctggc cttcgaggaa cagcctttcg
      901 atgtgatcgt ggcctacaga gtggaccggc tgaccagaag catccggcac ctgcaacagc
      961 tggtgcactg ggccgaggat cacaaaaagc tggtggtctc tgctacagag gcccacttcg
     1021 acacaaccac acctttcgcc gctgtggtga tcgccctgat ggggaccgtg gcccagatgg
     1081 aactcgaggc catcaaggaa cggaacagat ctgccgccca ctttaacatc agagccggca
     1141 agtacagagg aagcctgcca ccttggggct acctgcctac cagagttgac ggcgagtgga
     1201 gactggtccc cgaccccgtg cagagagagc ggatcctgga ggtgtaccac agagtggtcg
     1261 acaaccacga gcccctgcat ctggtggctc atgatctgaa tagaaggggc gtgctgagcc
     1321 ctaaggacta cttcgctcag ctgcagggca gagaacccca gggcagagag tggagcgcca
     1381 ccgccctgaa gagaagcatg atcagcgagg ccatgctggg atacgccacc ctgaacggca
     1441 aaaccgtgcg ggacgatgat ggagcccctc tggtgcgggc agagcctatc ctaacccgtg
     1501 agcagctgga agctctgaga gccgagctgg tcaagacatc tagagccaaa cctgctgtgt
     1561 ccacccctag cctgctgctg cgggtgctgt tctgtgccgt gtgcggtgaa cctgcttata
     1621 agttcgccgg cggcggcaga aagcacccca gataccggtg cagatctatg ggcttcccca
     1681 agcactgtgg caacggcacc gtggccatgg ccgagtggga tgctttttgc gaggaacaag
     1741 tgctggacct gctgggcgat gccgaaagac tagagaaggt gtgggtggct ggcagcgaca
     1801 gcgccgtgga actggctgaa gtgaacgccg agctggtgga cctcaccagc ctcatcggaa
     1861 gcccggccta ccgggccggc tctccacaga gagaagccct ggatgcccgt atcgccgccc
     1921 tggccgctag gcaggaggaa ctggaaggcc tggaagccag accaagcggc tgggagtgga
     1981 gagagacagg acagcggttc ggagattggt ggcgggaaca ggatacagcc gcaaagaaca
     2041 cctggctgag aagcatgaac gtgcggctga cattcgacgt gagaggcgga ctgaccagaa
     2101 caatcgactt cggcgacctg caggagtacg agcaacacct gagactgggc agcgtggtcg
     2161 agcggctgca cacaggcatg agctagtaat taattaagct gccttctgcg gggcttgcct
     2221 tctggccatg cccttcttct ctcccttgca cctgtacctc ttggtctttg aataaagcct
     2281 gagtaggaag cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
     2341 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgagaaaatt
     2401 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
     2461 aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct
     2521 tctgaggcgg aaagaaccag ctggggctcg ataccgtcga cctctagcta gagcttggcg
     2581 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     2641 atacgagccg gaagcataaa gtgtaaagcc tagggtgcct aatgagtgag ctaactcaca
     2701 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     2761 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     2821 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     2881 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     2941 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     3001 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     3061 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     3121 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     3181 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     3241 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     3301 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     3361 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     3421 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     3481 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     3541 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     3601 acggggtctg acactcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     3661 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     3721 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     3781 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     3841 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     3901 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     3961 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     4021 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     4081 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     4141 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     4201 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     4261 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     4321 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     4381 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4441 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     4501 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     4561 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     4621 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     4681 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     4741 gacgtcgacg gatcgggaga tcgatctccc gatcccctag ggtctactct cagtacaatc
     4801 tgctctgatg ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct
     4861 gagtagtgcg cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg
     4921 aagaatctgc ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg
     4981 cgtt
//
