LOCUS       sequence-205378-        6123 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  safe haven knock-in plasmid for the act5 locus, N-terminal GFP 
            tagging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6123)
  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 6123)
  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: pDM1513
FEATURES             Location/Qualifiers
     source          1..6123
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1503..1536
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1812..2849)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPDRE
                     MGEAN"
     protein_bind    3286..3319
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(4181..4198)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4352..4940)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4432..4451)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5111..5902,5903..5971))
                     /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     5734..5753
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5972..6076)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 actagcgccg gcattacatg tccatttatt agaaggattc aatcaaaaat taacaccagg
       61 tatattacca gatggcatta ataatttata ttctttaata ttggaaaaag aaaaaaataa
      121 ataaacaatc aatcagaaaa gataattgaa aaaaaaaaaa aagaaaatgt ttaataagaa
      181 ttaatttgga agtaaaataa tttttggatg agtaaatatt aataaactca caattaaaat
      241 ttagaatcca aaaaaatgaa aaaaaaaaaa atttttaaaa cagaatggtt gcattcattt
      301 tttttttttt ttttttatat cttttgggta tttttgggtg ttctatttat ctggataaaa
      361 aataaaatgg aatatagttt tatatatata tttttttcgt atcaagcttg aatgttttta
      421 tcaataatat gattgttggt tgctttattt tatttgaaaa aaaaattaat ttttaaaata
      481 tctttttttt cattttattt gaaaaaaaaa aaaaaaaaaa atggttttga aaatttttaa
      541 aaaaatggtt gggtcaccca caattcttat aaaaaaaagc catttggata gttttttttt
      601 tttttttttt tatttttttt attttttttt ttttgagatt tggtataaat acaaaaaaaa
      661 aattgttttt aaatcattat taataaaaac ttaaaataaa ttatatataa aaaaagatcc
      721 aaaaaatgtc aaaaggtgaa gaattattta caggtgttgt tccaatttta gttgaattag
      781 atggtgatgt taatggtcat aaattttcag tttcaggtga aggtgaaggt gatgcaacat
      841 atggtaaatt aacattaaaa tttatttgta caacaggtaa attaccagtt ccatggccaa
      901 cattagttac aacatttaca tatggtgttc aatgtttttc aagatatcca gatcatatga
      961 aacaacatga tttttttaaa tcagcaatgc cagaaggtta tgttcaagaa agaacaattt
     1021 tttttaaaga tgatggtaat tataaaacaa gagcagaagt taaatttgaa ggtgatacat
     1081 tagttaatag aattgaatta aaaggtattg attttaaaga agatggtaat attttaggtc
     1141 ataaattaga atataattat aattcacata atgtttatat tatggcagat aaacaaaaaa
     1201 atggtattaa agttaatttt aaaattagac ataatattga agatggttca gttcaattag
     1261 cagatcatta tcaacaaaat acaccaattg gtgatggtcc agttttatta ccagataatc
     1321 attatttatc aacacaatca gcattatcaa aagatccaaa tgaaaaaaga gatcatatgg
     1381 ttttattaga atttgttaca gcagcaggta ttacacatgg tatggatgaa ttatataaag
     1441 gtggttcagg aggtagtaga tcttcaacta gtataaaata aataaattgt ggatctgata
     1501 tcataacttc gtataatgta tgctatacga agttatacta ctctcgagat tttatttaat
     1561 atactaaata ataaaaaagt taaaaaatga tcattggata aattttttat aattataaat
     1621 aaagataata attttttttt taacaaaact aaaaataaaa ataataaaat aattgttaaa
     1681 ataggttttt tttttttttt ttttttttta ataaatggta tttattaatt tatttgttgt
     1741 gtgtgttttt ttttttataa tatttttttt tttagcattg aattaagaag aaatcaaatt
     1801 gatgcggccg cttagttagc ctcccccatc tcccgatccg gacgagtgct ggggcgtcgg
     1861 tttccactat cggcgagtac ttctacacag ccatcggtcc agacggccgc gcttctgcgg
     1921 gcgatttgtg tacgcccgac agtcccggct ccggatcgga cgattgcgtc gcatcgaccc
     1981 tgcgcccaag ctgcatcatc gaaattgccg tcaaccaagc tctgatagag ttggtcaaga
     2041 ccaatgcgga gcatatacgc ccggagccgc ggcgatcctg caagctccgg atgcctccgc
     2101 tcgaagtagc gcgtctgctg ctccatacaa gccaaccacg gcctccagaa gaagatgttg
     2161 gcgacctcgt attgggaatc cccgaacatc gcctcgctcc agtcaatgac cgctgttatg
     2221 cggccattgt ccgtcaggac attgttggag ccgaaatccg cgtgcacgag gtgccggact
     2281 tcggggcagt cctcggccca aagcatcagc tcatcgagag cctgcgcgac ggacgcactg
     2341 acggtgtcgt ccatcacagt ttgccagtga tacacatggg gatcagcaat cgcgcatatg
     2401 aaatcacgcc atgtagtgta ttgaccgatt ccttgcggtc cgaatgggcc gaacccgctc
     2461 gtctggctaa gatcggccgc agcgatcgca tccatggcct ccgcgaccgg ctgcagaaca
     2521 gcgggcagtt cggtttcagg caggtcttgc aacgtgacac cctgtgcacg gcgggagatg
     2581 caataggtca ggctctcgct gaactcccca atgtcaagca cttccggaat cgggagcgcg
     2641 gccgatgcaa agtgccgata aacataacga tctttgtaga aaccatcggc gcagctattt
     2701 acccgcagga catatccacg ccctcctaca tcgaagctga aagcacgaga ttcttcgccc
     2761 tccgagagct gcatcaggtc ggagacgctg tcgaactttt cgatcagaaa cttctcgaca
     2821 gacgtcgcgg tgagttcagg ctttttcatt aggtcttgtt gagaaatgtt aaattgatcc
     2881 attttttgct agctgtgaaa ttagtttaaa atacaaataa agagttataa taatatacag
     2941 ttgaataaaa aaaaaaaaaa tgaattggaa aatttatttt tatatgaaga aaaaaaaatt
     3001 ttgaaaaaaa aaaaaaaaat taaaaaaaaa aaaaaaaaaa aaaaaaatta aaaataattc
     3061 aactgtgggt gacccaaatt ttatttaaaa aaaaaaaaaa aatggatcac ttttgggtgg
     3121 ttggaaaaaa aaaaaaaaaa aaattgaaaa tataatgtta gtcatatgat taatcattaa
     3181 ttaattcttt tttttttttt ttttattttt tatttttttt ttttcaatat tccaattatg
     3241 acacacacta acatcacaaa atacaaaatt ttgggatcca ctactataac ttcgtataat
     3301 gtatgctata cgaagttatt taattaatta agatatcaag cttgtccaca gaaaatgttt
     3361 ctaaacaaaa aaaaaaaacc gagtgatgaa agcgcttctc acaaaattat ttatgtaaaa
     3421 tatttaataa aatacattat ttaaatcttt tttatttttt ttagttgttg atctttatcc
     3481 gactttaaaa ataaaaaaat tgtaaaaaaa aaaaaagttt atttgtttaa ttttattgtt
     3541 gtttttaatt ttttacaacc attaatttat aaaaccaaca cctaactata atgaaataca
     3601 atttgtatct tggaaaaaaa aaaattaata atataaataa aatgtaaaca agtatccggt
     3661 ggaaggaact aacagttgcc ttatccagtt caaaaaaaaa aaaaaaaaaa gaaattaaat
     3721 taccatttta tatttaacta aaataaatta aaaggtttat tttaataata gatttattat
     3781 tttttaaaaa tcaataatta atgttggata agatgaacga ataaaagata gagtttcagg
     3841 tcttttaaat gatgaggatt tatgaattct taaaagtttt gggcaagaga aaatgatttt
     3901 aacgattgaa ccatcactaa ttgcaaaatc tcttgaacta agagttttta agtttgaagc
     3961 aagtggagaa ttacaaagag caatcataga gttatcagtt aaatcataac aacaagataa
     4021 agataaagaa ctaatatgat gtaaattttc aatcatacct ctaaaagaaa catcattcat
     4081 actttttgga taaattaaat ctaatgaatc taaatttgga aatagtaaac caatatttgg
     4141 ccggcagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     4201 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     4261 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     4321 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     4381 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     4441 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     4501 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     4561 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     4621 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     4681 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     4741 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat
     4801 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     4861 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     4921 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     4981 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     5041 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     5101 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     5161 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     5221 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     5281 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5341 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5401 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     5461 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     5521 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     5581 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     5641 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     5701 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     5761 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     5821 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     5881 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     5941 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     6001 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     6061 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgcgccc tgtagcggga
     6121 tca
//
