LOCUS       sequence-490391-        4844 bp    DNA     circular SYN 23-OCT-2025
DEFINITION  Example plasmid for bridge recombinase mediated insertion and 
            off-target assessment with puromycin selection — must clone in UMI.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4844)
  AUTHORS   Perry NT, Bartie LJ, Katrekar D, Gonzalez GA, Durrant MG, Pai JJ, 
            Fanton A, Martins JQ, Hiraizumi M, Ricci-Tam C, Nishimasu H, 
            Konermann S, Hsu PD
  TITLE     Megabase-scale human genome rearrangement with programmable bridge 
            recombinases.
  JOURNAL   Science. 2025 Sep 25:eadz0276. doi: 10.1126/science.adz0276.
  PUBMED    40997214
REFERENCE   2  (bases 1 to 4844)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 23, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Science. 
            2025 Sep 25:eadz0276. doi: 10.1126/science.adz0276."
COMMENT     Sequence Label: pNP3386 [For use in genome insertion and off-t...
FEATURES             Location/Qualifiers
     source          1..4844
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..105
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(106..174,175..966)
                     /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(324..343)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1137..1725
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1626..1645
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1879..1896
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        2018..3196
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2249..3187
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3144..3164
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             3204..3800
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3204..3223)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3700..3720
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             3801..3857
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             3864..4574
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(4009..4027)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4277..4296
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(4599..4616)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4605..4812
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
       61 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
      121 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
      181 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
      241 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
      301 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
      361 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
      421 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
      481 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
      541 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
      601 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
      661 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
      721 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
      781 ggctggttta ttgctgataa atctggagcc ggtgagcgtg gctctcgcgg tatcattgca
      841 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
      901 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
      961 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     1021 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     1081 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     1141 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     1201 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     1261 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     1321 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     1381 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     1441 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     1501 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     1561 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     1621 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     1681 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     1741 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     1801 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     1861 agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg agcttcgcac
     1921 gattgcacct tgggagcacg gtacgtcggc tggacctcct aaatagttct aatccattca
     1981 ttgtgcattg ccgccgaatg caacctcgat cgagccgggc tccggtgccc gtcagtgggc
     2041 agagcgcaca tcgcccacag tccccgagaa gttgggggga ggggtcggca attgaaccgg
     2101 tgcctagaga aggtggcgcg gggtaaactg ggaaagtgat gtcgtgtact ggctccgcct
     2161 ttttcccgag ggtgggggag aaccgtatat aagtgcagta gtcgccgtga acgttctttt
     2221 tcgcaacggg tttgccgcca gaacacaggt aagtgccgtg tgtggttccc gcgggcctgg
     2281 cctctttacg ggttatggcc cttgcgtgcc ttgaattact tccacctggc tgcagtacgt
     2341 gattcttgat cccgagcttc gggttggaag tgggtgggag agttcgaggc cttgcgctta
     2401 aggagcccct tcgcctcgtg cttgagttga ggcctggcct gggcgctggg gccgccgcgt
     2461 gcgaatctgg tggcaccttc gcgcctgtct cgctgctttc gataagtctc tagccattta
     2521 aaatttttga tgacctgctg cgacgctttt tttctggcaa gatagtcttg taaatgcggg
     2581 ccaagatctg cacactggta tttcggtttt tggggccgcg ggcggcgacg gggcccgtgc
     2641 gtcccagcgc acatgttcgg cgaggcgggg cctgcgagcg cggccaccga gaatcggacg
     2701 ggggtagtct caagctggcc ggcctgctct ggtgcctggc ctcgcgccgc cgtgtatcgc
     2761 cccgccctgg gcggcaaggc tggcccggtc ggcaccagtt gcgtgagcgg aaagatggcc
     2821 gcttcccggc cctgctgcag ggagctcaaa atggaggacg cggcgctcgg gagagcgggc
     2881 gggtgagtca cccacacaaa ggaaaagggc ctttccgtcc tcagccgtcg cttcatgtga
     2941 ctccacggag taccgggcgc cgtccaggca cctcgattag ttctcgagct tttggagtac
     3001 gtcgtcttta ggttgggggg aggggtttta tgcgatggag tttccccaca ctgagtgggt
     3061 ggagactgaa gttaggccag cttggcactt gatgtaattc tccttggaat ttgccctttt
     3121 tgagtttgga tcttggttca ttctcaagcc tcagacagtg gttcaaagtt tttttcttcc
     3181 atttcaggtg tcgtgaagcc accatgaccg agtacaagcc cacggtgcgc ctcgccaccc
     3241 gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac taccccgcca
     3301 cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg caagaactct
     3361 tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac ggcgccgccg
     3421 tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc
     3481 cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg gaaggcctcc
     3541 tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcgga gtctcgcccg
     3601 accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag gcggccgagc
     3661 gcgccggggt gcccgccttc ctggaaacct ccgcgccccg caacctcccc ttctacgagc
     3721 ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc acctggtgca
     3781 tgacccgcaa gcccggtgcc gccacgaact tctctctgtt aaagcaagca ggtgacgtgg
     3841 aagaaaaccc cggtcccgga tccatggtga gcaagggcga ggaggataac atggccatca
     3901 tcaaggagtt catgcgcttc aaggtgcaca tggagggctc cgtgaacggc cacgagttcg
     3961 agatcgaggg cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg
     4021 tgaccaaggg tggccccctg cccttcgcct gggacatcct gtcccctcag ttcatgtacg
     4081 gctccaaggc ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc
     4141 ccgagggctt caagtgggag cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga
     4201 cccaggactc ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca
     4261 acttcccctc cgacggcccc gtaatgcaga agaagaccat gggctgggag gcctcctccg
     4321 agcggatgta ccccgaggac ggcgccctga agggcgagat caagcagagg ctgaagctga
     4381 aggacggcgg ccactacgac gctgaggtca agaccaccta caaggccaag aagcccgtgc
     4441 agctgcccgg cgcctacaac gtcaacatca agttggacat cacctcccac aacgaggact
     4501 acaccatcgt ggaacagtac gaacgcgccg agggccgcca ctccaccggc ggcatggacg
     4561 agctgtacaa gtaactcgag ctagagctcg ctgatcagcc tcgactgtgc cttctagttg
     4621 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     4681 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     4741 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     4801 gcatgctggg gacgacgtca ggtggcactt ttcggggaaa tgtg
//
