LOCUS       sequence-358228-        2969 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pDonor for ShHELIX containing 14bp of spacing between the I-AniI 
            site and LE/RE and encoding a kanamycin resistance gene as cargo.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2969)
  AUTHORS   Tou CJ, Orr B, Kleinstiver BP
  TITLE     Precise cut-and-paste DNA insertion using engineered type V-K 
            CRISPR-associated transposases.
  JOURNAL   Nat Biotechnol. 2023 Jan 2. doi: 10.1038/s41587-022-01574-x.
  PUBMED    36593413
REFERENCE   2  (bases 1 to 2969)
  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 
            Biotechnol. 2023 Jan 2. doi: 10.1038/s41587-022-01574-x."
COMMENT     Sequence Label: pDonor14-ShHELIX-KanR (CJT70)
FEATURES             Location/Qualifiers
     source          1..2969
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     oriT            184..292
                     /note="incP origin of transfer"
     protein_bind    complement(744..791)
                     /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             1178..1972
                     /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(1232..1251)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1842..1861
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     protein_bind    complement(2080..2127)
                     /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(2175..2261)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      complement(2297..2391)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      join(2746..2969,1..133)
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
ORIGIN
        1 aaacatgaga gcttagtacg ttagccatga gagcttagta cgttagccat gagggtttag
       61 ttcgttaaac atgagagctt agtacgttaa acttgagagc ttagtacgtg aaacatgaga
      121 gcttagtacg tactatcaac aggttgaact gcccatgttc tttcctgcgt tatcagagct
      181 tatcggccag cctcgcagag caggattccc gttgagcacc gccaggtgcg aataagggac
      241 agtgaagaag gaacacccgc tcgcgggtgg gcctacttca cctatcctgc ccggctgacg
      301 ccgttggata caccaaggaa agtctacacg aaccctttgg caaaatcctg tatatcgtgc
      361 gaaaaaggat ggatataccg aaaaaatcgc tataatgacc ccgaagcagg gttatgcagc
      421 ggaaagtagt actttagaca tctctttaca gagaaacctc ctcacacaaa aggcgtagtg
      481 tacagtgaca aattatctgt cgtcggtgac agattaatgt cattgtgact atttaattgt
      541 cgtcgtgacc catcagcgtt gcttaattaa ttgatgacaa attaaatgtc atcaatataa
      601 tatgctctgc aattattata caaagcaatt aaaacaagcg gataaaagga cttgctttca
      661 acccacccct aagtttaata gttactgagg gggatccact agtgagctca tgcatgatct
      721 cgaattagct tcaaaagcgc tctgaagttc ctatactttc tagagaatag gaacttcgga
      781 ataggaactt caagatcccc tgattccctt tgtcaacagc aatggataat tcgatttaac
      841 aaatgcatgg cgcaagggct gctaaaggaa gcggaacacg tagaaagcca gtccgcagaa
      901 acggtgctga ccccggatga atgtcagcta ctgggctatc tggacaaggg aaaacgcaag
      961 cgcaaagaga aagcaggtag cttgcagtgg gcttacatgg cgatagctag actgggcggt
     1021 tttatggaca gcaagcgaac cggaattgcc agctggggcg ccctctggta aggttgggaa
     1081 gccctgcaaa gtaaactgga tggctttctt gccgccaagg atctgatggc gcaggggatc
     1141 aagatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca
     1201 cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac
     1261 aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt
     1321 tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc
     1381 gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg
     1441 aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat cccaccttgc
     1501 tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc
     1561 ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat
     1621 ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc
     1681 cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca
     1741 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga
     1801 ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat
     1861 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc
     1921 tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct gaattgaaaa
     1981 aggaagagta tgaggatcca acatttccaa tcactagtga attatctaga attattccat
     2041 tgagtaagtt tttaagcaca tcagcttcaa aagcgctctg aagttcctat actttctaga
     2101 gaataggaac ttcggaatag gtacttcaag atccccaatt cgagatcgtc cgggccgcaa
     2161 gctcctagcg gcggatttgt cctactcagg agagcgttca ccgacaaaca acagataaaa
     2221 cgaaaggccc agtctttcga ctgagccttt cgttttattt gatgcctcaa gctagagagt
     2281 cattacccta gcttggattc tcaccaataa aaaacgcccg gcggcaaccg agcgttctga
     2341 acaaatccag atggagttct gaggtcatta ctggatctat caacaggagt ccaagctcag
     2401 ctaattaagg cgacagtcaa tttgtcatta tgaaaataca caaaagcttt ttcctatctt
     2461 gcaaagcgac agctaatttg tcacaatcac ggacaacgac atctattttg tcactgcaaa
     2521 gaggttatgc taaaactgcc aaagcgctat aatctatact gtataaggat tttactgatg
     2581 acaataattt gtcacaacga catataatta gtcactgtac acgtagagac gtagctgagg
     2641 aggtttctct gtaaaaatgc tacctctcgc gatatggtgc actctcagta caatcttgta
     2701 gcaccgccta catacctcgc tctgctaatc ctgttaccag ccggttgtca gccgttaagt
     2761 gttcctgtgt cactcaaaat tgctttgaga ggctctaagg gcttctcagt gcgttacatc
     2821 cctggcttgt tgtccacaac cgttaaacct taaaagcttt aaaagcctta tatattcttt
     2881 tttttcttat aaaacttaaa accttagagg ctatttaagt tgctgattta tattaatttt
     2941 attgttcaaa catgagagct tagtacgtg
//
