LOCUS       sequence-473906-        3646 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent reporter codon optimized for Dictyostelid species.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3646)
  TITLE     Selection-free CRISPR-Cas9 editing protocol for distant Dictyostelid
            species
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 3646)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pMG008_mCherry_P2A
FEATURES             Location/Qualifiers
     source          1..3646
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     278..300
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     292..309
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     293..309
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     320..339
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        320..338
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             396..416
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             468..515
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             516..581
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1287..1313
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1326..1382
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     primer_bind     complement(1789..1806)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1899..2487)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1979..1998)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2658..3449,3450..3518))
                     /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     3281..3300
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3519..3623)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtggccgct acagggcgct cccattcgcc attcaggctg cgcaactgtt
      181 gggaagggcg tttcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt
      241 gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg
      301 acggccagtg agcgcgacgt aatacgactc actatagggc gaattggcgg aaggccgtca
      361 aggccgcata aaaaatggtg agttcattaa gaccaccaaa gaaaaagaga aaggtttgtc
      421 caggtgatag atggagtagt acaggaggtg gaagaagtag aacttcaaag agacctgccg
      481 ctacaaaaaa ggccggacaa gcaaagaaga aaaaggacta caaagatcac gatggagatt
      541 ataaggatca cgatatagat tacaaagatg atgatgataa agtttcaaaa ggtgaagaag
      601 ataatatggc aattattaaa gaatttatga gatttaaagt tcatatggaa ggttcagtta
      661 atggtcatga atttgaaatt gaaggtgaag gtgaaggtag accatatgaa ggtacacaaa
      721 cagcaaaatt aaaagttaca aaaggtggtc cattaccatt tgcatgggat attttatcac
      781 cacaatttat gtatggttca aaagcatatg ttaaacatcc agcagatatt ccagattatt
      841 taaaattatc atttccagaa ggttttaaat gggaaagagt tatgaatttt gaagatggtg
      901 gtgttgttac agttacacaa gattcatcat tacaagatgg tgaatttatt tataaagtta
      961 aattaagagg tacaaatttt ccatcagatg gtccagttat gcaaaagaaa acaatgggtt
     1021 gggaagcatc atcagaaaga atgtatccag aagatggtgc attaaaaggt gaaattaaac
     1081 aaagattaaa attaaaagat ggtggtcatt atgatgcaga agttaaaaca acatataaag
     1141 caaaaaaacc agttcaatta ccaggtgcat ataatgttaa tattaaatta gatattacat
     1201 cacataatga agattataca attgttgaac aatatgaaag agcagaaggt agacattcaa
     1261 caggtggtat ggatgaatta tataaatacc catacgatgt acctgattat gctggtggtt
     1321 ctggtgctac aaatttctct ttgttaaagc aagctggaga tgttgaagaa aatcctggac
     1381 cttctagtgg acctagtgga tctagttctt gaattattta ataaataata aaaaaacaaa
     1441 ttgttgtaat aatctaatat tttctttttt ttttaatttt ttttttttaa atcttaataa
     1501 ttattaagtt attttaattt tttttttttt tttttttttt tttttttttt tttttctatc
     1561 aaaaaaatca aatatattta aaaaatttat tatttacaga tacattttga atggtgaaga
     1621 taaatatatg cattagatgt aaaacagcca aagagtatga aaatcaaaaa gatactgggc
     1681 ctcatgggcc ttccgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     1741 ttaacatggt catagctgtt tccttgcgta ttgggcgctc tccgcttcct cgctcactga
     1801 ctcgctgcgc tcggtcgttc gggtaaagcc tggggtgcct aatgagcaaa aggccagcaa
     1861 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     1921 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     1981 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     2041 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     2101 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     2161 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     2221 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     2281 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     2341 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     2401 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     2461 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     2521 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     2581 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     2641 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     2701 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     2761 ggcttaccat ctggccccag tgctgcaatg ataccgcgag aaccacgctc accggctcca
     2821 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     2881 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     2941 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     3001 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     3061 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     3121 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     3181 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     3241 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     3301 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     3361 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     3421 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     3481 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     3541 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     3601 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccac
//
