LOCUS       sequence-371123-        3553 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To chromosomally tag genes of interest with mCherry. 
            FRT-mCherry-cat-FRT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3553)
  AUTHORS   Avraham, Roi
  TITLE     Bacterial fluorescent reporters and tags toolkit
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 3553)
  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            "Unpublished"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: pKD3_mCherry
FEATURES             Location/Qualifiers
     source          1..3553
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(58..144)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     promoter        234..338
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(339..407,408..1199)
                     /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(557..576)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1301..1689
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     CDS             1701..2411
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGVYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1846..1864)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2114..2133
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    2449..2482
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     promoter        2588..2690
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             2691..3350
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPELRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2757..2776)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     protein_bind    3367..3414
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     terminator      complement(3492..3519)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 tcctgcccgc caccctccgg gccgttgctt cgcaacgttc aaatccgctc ccggcggatt
       61 tgtcctactc aggagagcgt tcaccgacaa acaacagata aaacgaaagg cccagtcttt
      121 cgactgagcc tttcatttta tttgatgcct ggcagttccc tactctcgca tggggagacc
      181 ccacactacc atcgggggcc atcgatgcag gtggcacttt tcggggaaat gtgcgcggaa
      241 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
      301 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
      361 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
      421 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
      481 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
      541 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
      601 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
      661 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
      721 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
      781 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
      841 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
      901 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
      961 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     1021 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     1081 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     1141 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     1201 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaatttt
     1261 tgcggccgca agatccggcc acgatgcgtc cggcgtagag gatctgaaga tcagcagttc
     1321 aacctgttga tagtacgtac taagctctca tgtttcacgt actaagctct catgtttaac
     1381 gtactaagct ctcatgttta acgaactaaa ccctcatggc taacgtacta agctctcatg
     1441 gctaacgtac taagctctca tgtttcacgt actaagctct catgtttgaa caataaaatt
     1501 aatataaatc agcaacttaa atagcctcta aggttttaag ttttataaga aaaaaaagaa
     1561 tatataaggc ttttaaagct tttaaggttt aacggttgtg gacaacaagc cagggatgta
     1621 acgcactgag aagcccttag agcctctcaa agcaattttg agtgacacag gaacacttaa
     1681 cggctgacat gggaattagc atggtgagca agggcgagga ggataacatg gccatcatca
     1741 aggagttcat gcgcttcaag gtgcacatgg agggctccgt gaacggccac gagttcgaga
     1801 tcgagggcga gggcgagggc cgcccctacg agggcaccca gaccgccaag ctgaaggtga
     1861 ccaagggtgg ccccctgccc ttcgcctggg acatcctgtc ccctcagttc atgtacggct
     1921 ccaaggccta cgtgaagcac cccgccgaca tccccgacta cttgaagctg tccttccccg
     1981 agggcttcaa gtgggagcgc gtgatgaact tcgaggacgg cggcgtggtg accgtgaccc
     2041 aggactcctc cctgcaggac ggcgagttca tctacaaggt gaagctgcgc ggcaccaact
     2101 tcccctccga cggccccgta atgcagaaga agaccatggg ctgggaggcc tcctccgagc
     2161 ggatgtaccc cgaggacggc gccctgaagg gcgagatcaa gcagaggctg aagctgaagg
     2221 acggcggcca ctacgacgct gaggtcaaga ccacctacaa ggccaagaag cccgtgcagc
     2281 tgcccggcgt ctacaacgtc aacatcaagt tggacatcac ctcccacaac gaggactaca
     2341 ccatcgtgga acagtacgaa cgcgccgagg gccgccactc caccggcggc atggacgagc
     2401 tgtacaagta acatggtcca tatgaatatc ctccttagtt cctattccga agttcctatt
     2461 ctctagaaag tataggaact tcggcgcgcc tacctgtgac ggaagatcac ttcgcagaat
     2521 aaataaatcc tggtgtccct gttgataccg ggaagccctg ggccaacttt tggcgaaaat
     2581 gagacgttga tcggcacgta agaggttcca actttcacca taatgaaata agatcactac
     2641 cgggcgtatt ttttgagttg tcgagatttt caggagctaa ggaagctaaa atggagaaaa
     2701 aaatcactgg atataccacc gttgatatat cccaatggca tcgtaaagaa cattttgagg
     2761 catttcagtc agttgctcaa tgtacctata accagaccgt tcagctggat attacggcct
     2821 ttttaaagac cgtaaagaaa aataagcaca agttttatcc ggcctttatt cacattcttg
     2881 cccgcctgat gaatgctcat ccggaattac gtatggcaat gaaagacggt gagctggtga
     2941 tatgggatag tgttcaccct tgttacaccg ttttccatga gcaaactgaa acgttttcat
     3001 cgctctggag tgaataccac gacgatttcc ggcagtttct acacatatat tcgcaagatg
     3061 tggcgtgtta cggtgaaaac ctggcctatt tccctaaagg gtttattgag aatatgtttt
     3121 tcgtctcagc caatccctgg gtgagtttca ccagttttga tttaaacgtg gccaatatgg
     3181 acaacttctt cgcccccgtt ttcaccatgg gcaaatatta tacgcaaggc gacaaggtgc
     3241 tgatgccgct ggcgattcag gttcatcatg ccgtttgtga tggcttccat gtcggcagaa
     3301 tgcttaatga attacaacag tactgcgatg agtggcaggg cggggcgtaa ggcgcgccat
     3361 ttaaatgaag ttcctattcc gaagttccta ttctctagaa agtataggaa cttcgaagca
     3421 gctccagcct acacaatcgc tcaagacgtg taatgctgca atctgcatgc aagcttggca
     3481 ctggccacgc aaaaaggcca tccgtcagga tggccttctg cttaatttga tgcctggcag
     3541 tttatggcgg gcg
//
