LOCUS       sequence-358229-        3026 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pDonor for AcHELIX containing 14bp of spacing between the I-AniI 
            site and LE/RE using ShCAST flanking sequence and encoding a 
            kanamycin resistance gene as cargo.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3026)
  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 3026)
  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-AcHELIX-KanR (CJT96)
FEATURES             Location/Qualifiers
     source          1..3026
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     oriT            252..360
                     /note="incP origin of transfer"
     protein_bind    complement(808..855)
                     /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             1242..2036
                     /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(1296..1315)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1906..1925
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     protein_bind    complement(2144..2191)
                     /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(2239..2325)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      complement(2428..2522)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     rep_origin      join(2871..3026,1..201)
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
ORIGIN
        1 ccttagaggc tatttaagtt gctgatttat attaatttta ttgttcaaac atgagagctt
       61 agtacgtgaa acatgagagc ttagtacgtt agccatgaga gcttagtacg ttagccatga
      121 gggtttagtt cgttaaacat gagagcttag tacgttaaac ttgagagctt agtacgtgaa
      181 acatgagagc ttagtacgta ctatcaacag gttgaactgc ccatgttctt tcctgcgtta
      241 tcagagctta tcggccagcc tcgcagagca ggattcccgt tgagcaccgc caggtgcgaa
      301 taagggacag tgaagaagga acacccgctc gcgggtgggc ctacttcacc tatcctgccc
      361 ggctgacgcc gttggataca ccaaggaaag tctacacgaa ccctttggca aaatcctgta
      421 tatcgtgcga aaaaggatgg atataccgaa aaaatcgcta taatgacccc gaagcagggt
      481 tatgcagcgg aaagtagtac tttagacatc tctttacaga gaaacctcct cacacaaaag
      541 gcgtagtgta cattcgcaaa ttaaatgtcg cttttcgcaa tttagtgtcg ttattcgcaa
      601 attaatgtcg tggtggttgt ttttcagagt caatttaatt attctaagtt ttcgcaaatt
      661 aatgtcgcat gaacttaaca tttactatac aataaattat tgctgcaagg gcattattgg
      721 attattgata tgtgttcgat cgcagcactc ctgggggatc cactagtgag ctcatgcatg
      781 atctcgaatt agcttcaaaa gcgctctgaa gttcctatac tttctagaga ataggaactt
      841 cggaatagga acttcaagat cccctgattc cctttgtcaa cagcaatgga taattcgatt
      901 taacaaatgc atggcgcaag ggctgctaaa ggaagcggaa cacgtagaaa gccagtccgc
      961 agaaacggtg ctgaccccgg atgaatgtca gctactgggc tatctggaca agggaaaacg
     1021 caagcgcaaa gagaaagcag gtagcttgca gtgggcttac atggcgatag ctagactggg
     1081 cggttttatg gacagcaagc gaaccggaat tgccagctgg ggcgccctct ggtaaggttg
     1141 ggaagccctg caaagtaaac tggatggctt tcttgccgcc aaggatctga tggcgcaggg
     1201 gatcaagatc tgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat
     1261 tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac
     1321 agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc
     1381 tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaggacgag gcagcgcggc
     1441 tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag
     1501 cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatcccacc
     1561 ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg
     1621 atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc
     1681 ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc
     1741 cagccgaact gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga
     1801 cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca
     1861 tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg
     1921 atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg
     1981 ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgaattg
     2041 aaaaaggaag agtatgagga tccaacattt ccaatcacta gtgaattatc tagaattatt
     2101 ccattgagta agtttttaag cacatcagct tcaaaagcgc tctgaagttc ctatactttc
     2161 tagagaatag gaacttcgga ataggtactt caagatcccc aattcgagat cgtccgggcc
     2221 gcaagctcct agcggcggat ttgtcctact caggagagcg ttcaccgaca aacaacagat
     2281 aaaacgaaag gcccagtctt tcgactgagc ctttcgtttt atttgatgcc tcaagctaga
     2341 gagtcattac cccaggcgtt taagggcacc aataactgcc ttaaaaaaat tacgccccgc
     2401 cctgccactc atcgcagtct agcttggatt ctcaccaata aaaaacgccc ggcggcaacc
     2461 gagcgttctg aacaaatcca gatggagttc tgaggtcatt actggatcta tcaacaggag
     2521 tccaagctca gctaattaag gacatctaat ttgcaaaata ccaaattctt aacaaacgac
     2581 atttaatttg cgaaaccagg ttttacgaca tacaatatgc gaattaggta acttagtctt
     2641 ttgtaggggt aaatagctta tgatgcttat agaataaagg ttttagtcct taaaagcagt
     2701 tgcgacacta atttgcgaaa agcgacattt aatttgcgaa cgtacacgta gagacgtagc
     2761 tgaggaggtt tctctgtaaa aatgctacct ctcgcgatat ggtgcactct cagtacaatc
     2821 ttgtagcacc gcctacatac ctcgctctgc taatcctgtt accagccggt tgtcagccgt
     2881 taagtgttcc tgtgtcactc aaaattgctt tgagaggctc taagggcttc tcagtgcgtt
     2941 acatccctgg cttgttgtcc acaaccgtta aaccttaaaa gctttaaaag ccttatatat
     3001 tctttttttt cttataaaac ttaaaa
//
