LOCUS       sequence-248516-        3005 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Kanamycin resistance donor for AcCAST.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3005)
  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 3005)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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_AcCAST_kanR
FEATURES             Location/Qualifiers
     source          1..3005
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     oriT            19..127
                     /note="incP origin of transfer"
     protein_bind    complement(565..612)
                     /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             999..1793
                     /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(1053..1072)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1663..1682
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     protein_bind    complement(1901..1948)
                     /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(1996..2082)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      complement(2185..2279)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      2617..2973
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
ORIGIN
        1 tgcgttatca gagcttatcg gccagcctcg cagagcagga ttcccgttga gcaccgccag
       61 gtgcgaataa gggacagtga agaaggaaca cccgctcgcg ggtgggccta cttcacctat
      121 cctgcccggc tgacgccgtt ggatacacca aggaaagtct acacgaaccc tttggcaaaa
      181 tcctgtatat cgtgcgaaaa aggatggata taccgaaaaa atcgctataa tgaccccgaa
      241 gcagggttat gcagcggaaa gtacagaggt caggggttcg agtctcctat tctccattat
      301 atatgtacat tcgcaaatta aatgtcgctt ttcgcaattt agtgtcgtta ttcgcaaatt
      361 aatgtcgtgg tggttgtttt tcagagtcaa tttaattatt ctaagttttc gcaaattaat
      421 gtcgcatgaa cttaacattt actatacaat aaattattgc tgcaagggca ttattggatt
      481 attgatatgt gttcgatcgc agcactcctg ggggatccac tagtgagctc atgcatgatc
      541 tcgaattagc ttcaaaagcg ctctgaagtt cctatacttt ctagagaata ggaacttcgg
      601 aataggaact tcaagatccc ctgattccct ttgtcaacag caatggataa ttcgatttaa
      661 caaatgcatg gcgcaagggc tgctaaagga agcggaacac gtagaaagcc agtccgcaga
      721 aacggtgctg accccggatg aatgtcagct actgggctat ctggacaagg gaaaacgcaa
      781 gcgcaaagag aaagcaggta gcttgcagtg ggcttacatg gcgatagcta gactgggcgg
      841 ttttatggac agcaagcgaa ccggaattgc cagctggggc gccctctggt aaggttggga
      901 agccctgcaa agtaaactgg atggctttct tgccgccaag gatctgatgg cgcaggggat
      961 caagatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
     1021 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     1081 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     1141 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat
     1201 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     1261 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tcccaccttg
     1321 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     1381 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     1441 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     1501 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc
     1561 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     1621 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     1681 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     1741 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgaattgaaa
     1801 aaggaagagt atgaggatcc aacatttcca atcactagtg aattatctag aattattcca
     1861 ttgagtaagt ttttaagcac atcagcttca aaagcgctct gaagttccta tactttctag
     1921 agaataggaa cttcggaata ggtacttcaa gatccccaat tcgagatcgt ccgggccgca
     1981 agctcctagc ggcggatttg tcctactcag gagagcgttc accgacaaac aacagataaa
     2041 acgaaaggcc cagtctttcg actgagcctt tcgttttatt tgatgcctca agctagagag
     2101 tcattacccc aggcgtttaa gggcaccaat aactgcctta aaaaaattac gccccgccct
     2161 gccactcatc gcagtctagc ttggattctc accaataaaa aacgcccggc ggcaaccgag
     2221 cgttctgaac aaatccagat ggagttctga ggtcattact ggatctatca acaggagtcc
     2281 aagctcagct aattaaggac atctaatttg caaaatacca aattcttaac aaacgacatt
     2341 taatttgcga aaccaggttt tacgacatac aatatgcgaa ttaggtaact tagtcttttg
     2401 taggggtaaa tagcttatga tgcttataga ataaaggttt tagtccttaa aagcagttgc
     2461 gacactaatt tgcgaaaagc gacatttaat ttgcgaacgt acaatagcct ttctcactct
     2521 agttagatac aaaattaccc ctctatggtg cactctcagt acaatcttgt agcaccgcct
     2581 acatacctcg ctctgctaat cctgttacca gccggttgtc agccgttaag tgttcctgtg
     2641 tcactcaaaa ttgctttgag aggctctaag ggcttctcag tgcgttacat ccctggcttg
     2701 ttgtccacaa ccgttaaacc ttaaaagctt taaaagcctt atatattctt ttttttctta
     2761 taaaacttaa aaccttagag gctatttaag ttgctgattt atattaattt tattgttcaa
     2821 acatgagagc ttagtacgtg aaacatgaga gcttagtacg ttagccatga gagcttagta
     2881 cgttagccat gagggtttag ttcgttaaac atgagagctt agtacgttaa acttgagagc
     2941 ttagtacgtg aaacatgaga gcttagtacg tactatcaac aggttgaact gcccatgttc
     3001 tttcc
//
