LOCUS       sequence-414393-        4564 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Knock in plasmid for mNeonGreen fusions C-terminal .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4564)
  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 4564)
  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: pPI462
FEATURES             Location/Qualifiers
     source          1..4564
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    723..756
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1030..2067)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPDRE
                     MGEAN"
     protein_bind    2503..2536
                     /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(2622..2639)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2793..3381)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2873..2892)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3552..4343,4344..4412))
                     /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     4175..4194
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4413..4517)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 actagtggtg gttcaggagg tagtgtttca aaaggtgaag aagataatat ggcatcgtta
       61 ccagctacac atgagttaca tatattcggt agcattaatg gtgttgattt tgatatggtg
      121 ggacaaggta ccggtaatcc taatgatggt tacgaagaac taaatttaaa atcgactaaa
      181 ggtgacttac aattttctcc atggatttta gtgccacata tagggtatgg ttttcatcaa
      241 tacttaccat atccagatgg tatgtcacca tttcaagctg caatggttga tggatcaggt
      301 tatcaagttc atagaacaat gcaatttgaa gatggtgctt cattaactgt taattataga
      361 tacacatatg aaggctcaca tattaaaggt gaagctcaag ttaaaggtac tggtttccca
      421 gccgatggcc cagttatgac aaatagttta acagcagcag attggtgtag atccaaaaaa
      481 acttatccaa atgataaaac aattatttca acttttaaat ggtcatatac aaccggtaat
      541 ggtaaacgtt atcgttcaac agcccgtaca acatatactt ttgctaaacc aatggcagct
      601 aattatttaa aaaatcaacc aatgtatgtt tttcgtaaaa cagagttaaa acattcaaaa
      661 acagaactta attttaaaga atggcaaaaa gcatttacag acgttatgta agatctgata
      721 tcataacttc gtataatgta tgctatacga agttatacta ctctcgagat tttatttaat
      781 atactaaata ataaaaaagt taaaaaatga tcattggata aattttttat aattataaat
      841 aaagataata attttttttt taacaaaact aaaaataaaa ataataaaat aattgttaaa
      901 ataggttttt tttttttttt ttttttttaa taaatggtat ttattaattt atttgttgtg
      961 tgtgtttttt ttttataata tttttttttt tagcattgaa ttaagaagaa atcaaattga
     1021 tgcggccgct tagttagcct cccccatctc ccgatccgga cgagtgctgg ggcgtcggtt
     1081 tccactatcg gcgagtactt ctacacagcc atcggtccag acggccgcgc ttctgcgggc
     1141 gatttgtgta cgcccgacag tcccggctcc ggatcggacg attgcgtcgc atcgaccctg
     1201 cgcccaagct gcatcatcga aattgccgtc aaccaagctc tgatagagtt ggtcaagacc
     1261 aatgcggagc atatacgccc ggagccgcgg cgatcctgca agctccggat gcctccgctc
     1321 gaagtagcgc gtctgctgct ccatacaagc caaccacggc ctccagaaga agatgttggc
     1381 gacctcgtat tgggaatccc cgaacatcgc ctcgctccag tcaatgaccg ctgttatgcg
     1441 gccattgtcc gtcaggacat tgttggagcc gaaatccgcg tgcacgaggt gccggacttc
     1501 ggggcagtcc tcggcccaaa gcatcagctc atcgagagcc tgcgcgacgg acgcactgac
     1561 ggtgtcgtcc atcacagttt gccagtgata cacatgggga tcagcaatcg cgcatatgaa
     1621 atcacgccat gtagtgtatt gaccgattcc ttgcggtccg aatgggccga acccgctcgt
     1681 ctggctaaga tcggccgcag cgatcgcatc catggcctcc gcgaccggct gcagaacagc
     1741 gggcagttcg gtttcaggca ggtcttgcaa cgtgacaccc tgtgcacggc gggagatgca
     1801 ataggtcagg ctctcgctga actccccaat gtcaagcact tccggaatcg ggagcgcggc
     1861 cgatgcaaag tgccgataaa cataacgatc tttgtagaaa ccatcggcgc agctatttac
     1921 ccgcaggaca tatccacgcc ctcctacatc gaagctgaaa gcacgagatt cttcgccctc
     1981 cgagagctgc atcaggtcgg agacgctgtc gaacttttcg atcagaaact tctcgacaga
     2041 cgtcgcggtg agttcaggct ttttcattag gtcttgttga gaaatgttaa attgatccat
     2101 tttttgctag ctgtgaaatt agtttaaaat acaaataaag agttataata atatacagtt
     2161 gaataaaaaa aaaaaaaawr aattgraaaa tttattttta wawaaaaaaa aaaaaawttt
     2221 aaaaaaaaaa aaaaaaawwa aaaaaaaaaa aaaaaaaaaa aaaaaaaaaa aaaaattmaa
     2281 ctgtgggtga cccaaatttt atttaaaaaa aaaaaaaaaa tggatcactt ttgggtggtt
     2341 ggaaaaaaaa aaaaaaaaaa attgaaaata taatgttagt catatgatta atcattaatt
     2401 aattcttttt tttttttttt ttatttttta tttttttttt tcaatattcc aattatgaca
     2461 cacactaaca tcacaaaata caaaattttg ggatccacta ctataacttc gtataatgta
     2521 tgctatacga agttatttaa ttaattaaga tatcaagctt gatagctctg cctactgaag
     2581 gccggcagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
     2641 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     2701 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     2761 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     2821 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     2881 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     2941 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     3001 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     3061 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     3121 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     3181 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     3241 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     3301 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     3361 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     3421 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     3481 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     3541 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     3601 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     3661 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     3721 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     3781 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     3841 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     3901 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     3961 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     4021 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     4081 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     4141 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     4201 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     4261 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     4321 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     4381 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     4441 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     4501 caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgcgcc ctgtagcggg
     4561 atca
//
