LOCUS       sequence-205379-        6117 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  safe haven knock-in plasmid for the act5 locus, N-terminal mCherry 
            tagging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6117)
  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 6117)
  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: pDM1514
FEATURES             Location/Qualifiers
     source          1..6117
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1497..1530
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1806..2843)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPDRE
                     MGEAN"
     protein_bind    3280..3313
                     /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(4175..4192)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4346..4934)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4426..4445)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5105..5896,5897..5965))
                     /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     5728..5747
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5966..6070)
                     /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 aaaaaatggt ttcaaaaggt gaagaagata atatggcaat tattaaagaa tttatgagat
      781 ttaaagttca tatggaaggt tcagttaatg gtcatgaatt tgaaattgaa ggtgaaggtg
      841 aaggtagacc atatgaaggt acacaaacag caaaattaaa agttacaaaa ggtggtccat
      901 taccatttgc atgggatatt ttatcaccac aatttatgta tggttcaaaa gcatatgtta
      961 aacatccagc agatattcca gattatttaa aattatcatt tccagaaggt tttaaatggg
     1021 aaagagttat gaattttgaa gatggtggtg ttgttacagt tacacaagat tcatcattac
     1081 aagatggtga atttatttat aaagttaaat taagaggtac aaattttcca tcagatggtc
     1141 cagttatgca aaaaaaaaca atgggttggg aagcatcatc agaaagaatg tatccagaag
     1201 atggtgcatt aaaaggtgaa attaaacaaa gattaaaatt aaaagatggt ggtcattatg
     1261 atgcagaagt taaaacaaca tataaagcaa aaaaaccagt tcaattacca ggtgcatata
     1321 atgttaatat taaattagat attacatcac ataatgaaga ttatacaatt gttgaacaat
     1381 atgaaagagc agaaggtaga cattcaacag gtggtatgga tgaattatat aaaggtggtt
     1441 caggaggtag tagatcttca actagtataa aataaataaa ttgtggatct gatatcataa
     1501 cttcgtataa tgtatgctat acgaagttat actactctcg agattttatt taatatacta
     1561 aataataaaa aagttaaaaa atgatcattg gataaatttt ttataattat aaataaagat
     1621 aataattttt tttttaacaa aactaaaaat aaaaataata aaataattgt taaaataggt
     1681 tttttttttt tttttttttt tttaataaat ggtatttatt aatttatttg ttgtgtgtgt
     1741 tttttttttt ataatatttt tttttttagc attgaattaa gaagaaatca aattgatgcg
     1801 gccgcttagt tagcctcccc catctcccga tccggacgag tgctggggcg tcggtttcca
     1861 ctatcggcga gtacttctac acagccatcg gtccagacgg ccgcgcttct gcgggcgatt
     1921 tgtgtacgcc cgacagtccc ggctccggat cggacgattg cgtcgcatcg accctgcgcc
     1981 caagctgcat catcgaaatt gccgtcaacc aagctctgat agagttggtc aagaccaatg
     2041 cggagcatat acgcccggag ccgcggcgat cctgcaagct ccggatgcct ccgctcgaag
     2101 tagcgcgtct gctgctccat acaagccaac cacggcctcc agaagaagat gttggcgacc
     2161 tcgtattggg aatccccgaa catcgcctcg ctccagtcaa tgaccgctgt tatgcggcca
     2221 ttgtccgtca ggacattgtt ggagccgaaa tccgcgtgca cgaggtgccg gacttcgggg
     2281 cagtcctcgg cccaaagcat cagctcatcg agagcctgcg cgacggacgc actgacggtg
     2341 tcgtccatca cagtttgcca gtgatacaca tggggatcag caatcgcgca tatgaaatca
     2401 cgccatgtag tgtattgacc gattccttgc ggtccgaatg ggccgaaccc gctcgtctgg
     2461 ctaagatcgg ccgcagcgat cgcatccatg gcctccgcga ccggctgcag aacagcgggc
     2521 agttcggttt caggcaggtc ttgcaacgtg acaccctgtg cacggcggga gatgcaatag
     2581 gtcaggctct cgctgaactc cccaatgtca agcacttccg gaatcgggag cgcggccgat
     2641 gcaaagtgcc gataaacata acgatctttg tagaaaccat cggcgcagct atttacccgc
     2701 aggacatatc cacgccctcc tacatcgaag ctgaaagcac gagattcttc gccctccgag
     2761 agctgcatca ggtcggagac gctgtcgaac ttttcgatca gaaacttctc gacagacgtc
     2821 gcggtgagtt caggcttttt cattaggtct tgttgagaaa tgttaaattg atccattttt
     2881 tgctagctgt gaaattagtt taaaatacaa ataaagagtt ataataatat acagttgaat
     2941 aaaaaaaaaa aaaatgaatt ggaaaattta tttttatatg aagaaaaaaa aattttgaaa
     3001 aaaaaaaaaa aaattaaaaa aaaaaaaaaa aaaaaaaaaa attaaaaata attcaactgt
     3061 gggtgaccca aattttattt aaaaaaaaaa aaaaaatgga tcacttttgg gtggttggaa
     3121 aaaaaaaaaa aaaaaaattg aaaatataat gttagtcata tgattaatca ttaattaatt
     3181 cttttttttt ttttttttat tttttatttt ttttttttca atattccaat tatgacacac
     3241 actaacatca caaaatacaa aattttggga tccactacta taacttcgta taatgtatgc
     3301 tatacgaagt tatttaatta attaagatat caagcttgtc cacagaaaat gtttctaaac
     3361 aaaaaaaaaa aaccgagtga tgaaagcgct tctcacaaaa ttatttatgt aaaatattta
     3421 ataaaataca ttatttaaat cttttttatt ttttttagtt gttgatcttt atccgacttt
     3481 aaaaataaaa aaattgtaaa aaaaaaaaaa gtttatttgt ttaattttat tgttgttttt
     3541 aattttttac aaccattaat ttataaaacc aacacctaac tataatgaaa tacaatttgt
     3601 atcttggaaa aaaaaaaatt aataatataa ataaaatgta aacaagtatc cggtggaagg
     3661 aactaacagt tgccttatcc agttcaaaaa aaaaaaaaaa aaaagaaatt aaattaccat
     3721 tttatattta actaaaataa attaaaaggt ttattttaat aatagattta ttatttttta
     3781 aaaatcaata attaatgttg gataagatga acgaataaaa gatagagttt caggtctttt
     3841 aaatgatgag gatttatgaa ttcttaaaag ttttgggcaa gagaaaatga ttttaacgat
     3901 tgaaccatca ctaattgcaa aatctcttga actaagagtt tttaagtttg aagcaagtgg
     3961 agaattacaa agagcaatca tagagttatc agttaaatca taacaacaag ataaagataa
     4021 agaactaata tgatgtaaat tttcaatcat acctctaaaa gaaacatcat tcatactttt
     4081 tggataaatt aaatctaatg aatctaaatt tggaaatagt aaaccaatat ttggccggca
     4141 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     4201 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     4261 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     4321 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     4381 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     4441 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     4501 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     4561 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     4621 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     4681 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     4741 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     4801 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     4861 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     4921 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     4981 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     5041 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     5101 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     5161 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     5221 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     5281 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     5341 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     5401 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     5461 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     5521 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     5581 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     5641 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     5701 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     5761 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     5821 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     5881 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     5941 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     6001 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     6061 gggttccgcg cacatttccc cgaaaagtgc cacctgacgc gccctgtagc gggatca
//
