LOCUS       sequence-205362-        3643 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Knockout cassette G418.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3643)
  AUTHORS   Paschke P, Knecht DA, Silale A, Traynor D, Williams TD, Thomason PA,
            Insall RH, Chubb JR, Kay RR, Veltman DM
  TITLE     Rapid and efficient genetic engineering of both wild type and axenic
            strains of Dictyostelium discoideum.
  JOURNAL   PLoS One. 2018 May 30;13(5):e0196809. doi: 
            10.1371/journal.pone.0196809. eCollection 2018.
  PUBMED    29847546
REFERENCE   2  (bases 1 to 3643)
  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: "PLoS One.";
            date: "2018-05-30"; volume: "13(5):e0196809. doi"; pages: " 
            10.1371/journal.pone.0196809. eCollection 2018"
COMMENT     Sequence Label: pDM1082
FEATURES             Location/Qualifiers
     source          1..3643
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(51..84)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             524..1318
                     /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(578..597)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1188..1207
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     protein_bind    complement(1594..1627)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        1713..1817
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1818..1886,1887..2678)
                     /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(2036..2055)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2849..3437
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3338..3357
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3591..3608
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gccggccttc agtaggcaga gctatcaagc ttgatatctt aattaattaa ataacttcgt
       61 atagcataca ttatacgaag ttatagtagt ggatcccaaa attttgtatt ttgtgatgtt
      121 agtgtgtgtc ataattggaa tattgaaaaa aaaaaaataa aaaataaaaa aaaaaaaaaa
      181 aagaattaat taatgattaa tcatatgact aacattatat tttcaatttt tttttttttt
      241 tttttccaac cacccaaaag tgatccattt tttttttttt tttaaataaa atttgggtca
      301 cccacagttg aattattttt aatttttttt tttttttttt ttttttttaa tttttttttt
      361 ttttttcaaa attttttttt cttcatataa aaataaattt tccaattcat tttttttttt
      421 tttattcaac tgtatattat tataactctt tatttgtatt ttaaactaat ttcacagcta
      481 gcaaaaaatg gatggtgaag atgttcaagc tggatcgttt cgcatgattg aacaagatgg
      541 attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca
      601 acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt
      661 tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg
      721 gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga
      781 agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca
      841 ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct
      901 tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac
      961 tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc
     1021 gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt
     1081 gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt
     1141 catcgactgt ggacggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg
     1201 tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat
     1261 cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc
     1321 ggccgcatca atttgatttc ttcttaattc aatgctaaaa aaaaaaatat tataaaaaaa
     1381 aaaacacaca caacaaataa attaataaat accatttatt aaaaaaaaaa aaaaaaaaaa
     1441 aaaacctatt ttaacaatta ttttattatt tttattttta gttttgttaa aaaaaaaatt
     1501 attatcttta tttataatta taaaaaattt atccaatgat cattttttaa cttttttatt
     1561 atttagtata ttaaataaaa tctcgagagt agtataactt cgtatagcat acattatacg
     1621 aagttatgat atcagatctc tactggagta tccaagctga ctagttgatc ccgctacagg
     1681 gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     1741 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     1801 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     1861 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     1921 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     1981 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
     2041 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact
     2101 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     2161 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     2221 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     2281 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     2341 agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg
     2401 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
     2461 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
     2521 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
     2581 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
     2641 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
     2701 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     2761 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     2821 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     2881 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     2941 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc
     3001 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     3061 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     3121 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     3181 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     3241 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     3301 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     3361 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     3421 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     3481 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     3541 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
     3601 tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tct
//
