LOCUS       sequence-245550-        2999 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Kanamycin resistance donor for ShCAST.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2999)
  AUTHORS   Strecker J, Ladha A, Gardner Z, Schmid-Burgk JL, Makarova KS, Koonin
            EV, Zhang F
  TITLE     RNA-guided DNA insertion with CRISPR-associated transposases.
  JOURNAL   Science. 2019 Jul 5;365(6448):48-53. doi: 10.1126/science.aax9181. 
            Epub 2019 Jun 6.
  PUBMED    31171706
REFERENCE   2  (bases 1 to 2999)
  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"; doi: 
            "10.1126/science.aax9181"; journalName: "Science"; date: "2019-07-5-
            5"; volume: "365"; issue: "6448"; pages: "48-53"
COMMENT     Sequence Label: pDonor_ShCAST_kanR
FEATURES             Location/Qualifiers
     source          1..2999
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..357
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     oriT            408..516
                     /note="incP origin of transfer"
     protein_bind    complement(959..1006)
                     /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)."
     CDS             1393..2187
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1447..1466)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2057..2076
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     protein_bind    complement(2295..2342)
                     /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)."
     terminator      complement(2390..2476)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      complement(2579..2673)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
ORIGIN
        1 tgtcagccgt taagtgttcc tgtgtcactc aaaattgctt tgagaggctc taagggcttc
       61 tcagtgcgtt acatccctgg cttgttgtcc acaaccgtta aaccttaaaa gctttaaaag
      121 ccttatatat tctttttttt cttataaaac ttaaaacctt agaggctatt taagttgctg
      181 atttatatta attttattgt tcaaacatga gagcttagta cgtgaaacat gagagcttag
      241 tacgttagcc atgagagctt agtacgttag ccatgagggt ttagttcgtt aaacatgaga
      301 gcttagtacg ttaaacttga gagcttagta cgtgaaacat gagagcttag tacgtactat
      361 caacaggttg aactgcccat gttctttcct gcgttatcag agcttatcgg ccagcctcgc
      421 agagcaggat tcccgttgag caccgccagg tgcgaataag ggacagtgaa gaaggaacac
      481 ccgctcgcgg gtgggcctac ttcacctatc ctgcccggct gacgccgttg gatacaccaa
      541 ggaaagtcta cacgaaccct ttggcaaaat cctgtatatc gtgcgaaaaa ggatggatat
      601 accgaaaaaa tcgctataat gaccccgaag cagggttatg cagcggaaag taaaaaattt
      661 ttagtttatt agacatctcc acaaaaggcg tagtgtacag tgacaaatta tctgtcgtcg
      721 gtgacagatt aatgtcattg tgactattta attgtcgtcg tgacccatca gcgttgctta
      781 attaattgat gacaaattaa atgtcatcaa tataatatgc tctgcaatta ttatacaaag
      841 caattaaaac aagcggataa aaggacttgc tttcaaccca cccctaagtt taatagttac
      901 tgagggggat ccactagtga gctcatgcat gatctcgaat tagcttcaaa agcgctctga
      961 agttcctata ctttctagag aataggaact tcggaatagg aacttcaaga tcccctgatt
     1021 ccctttgtca acagcaatgg ataattcgat ttaacaaatg catggcgcaa gggctgctaa
     1081 aggaagcgga acacgtagaa agccagtccg cagaaacggt gctgaccccg gatgaatgtc
     1141 agctactggg ctatctggac aagggaaaac gcaagcgcaa agagaaagca ggtagcttgc
     1201 agtgggctta catggcgata gctagactgg gcggttttat ggacagcaag cgaaccggaa
     1261 ttgccagctg gggcgccctc tggtaaggtt gggaagccct gcaaagtaaa ctggatggct
     1321 ttcttgccgc caaggatctg atggcgcagg ggatcaagat ctgatcaaga gacaggatga
     1381 ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg
     1441 gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg
     1501 ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc
     1561 ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct
     1621 tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa
     1681 gtgccggggc aggatctcct gtcatcccac cttgctcctg ccgagaaagt atccatcatg
     1741 gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa
     1801 gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat
     1861 gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg
     1921 cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc
     1981 atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac
     2041 cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg
     2101 gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc
     2161 tatcgccttc ttgacgagtt cttctgaatt gaaaaaggaa gagtatgagg atccaacatt
     2221 tccaatcact agtgaattat ctagaattat tccattgagt aagtttttaa gcacatcagc
     2281 ttcaaaagcg ctctgaagtt cctatacttt ctagagaata ggaacttcgg aataggtact
     2341 tcaagatccc caattcgaga tcgtccgggc cgcaagctcc tagcggcgga tttgtcctac
     2401 tcaggagagc gttcaccgac aaacaacaga taaaacgaaa ggcccagtct ttcgactgag
     2461 cctttcgttt tatttgatgc ctcaagctag agagtcatta ccccaggcgt ttaagggcac
     2521 caataactgc cttaaaaaaa ttacgccccg ccctgccact catcgcagtc tagcttggat
     2581 tctcaccaat aaaaaacgcc cggcggcaac cgagcgttct gaacaaatcc agatggagtt
     2641 ctgaggtcat tactggatct atcaacagga gtccaagctc agctaattaa ggcgacagtc
     2701 aatttgtcat tatgaaaata cacaaaagct ttttcctatc ttgcaaagcg acagctaatt
     2761 tgtcacaatc acggacaacg acatctattt tgtcactgca aagaggttat gctaaaactg
     2821 ccaaagcgct ataatctata ctgtataagg attttactga tgacaataat ttgtcacaac
     2881 gacatataat tagtcactgt acacgtagag acgtagcaat gctacctctc tacaatggtt
     2941 ttgtatggtg cactctcagt acaatctgcc tcgctctgct aatcctgtta ccagccggt
//
