LOCUS       sequence-371233-        7591 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses nuclear-localized mCherry-XL (R125K) in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7591)
  AUTHORS   Mukherjee S, Manna P, Hung ST, Vietmeyer F, Friis P, Palmer AE, 
            Jimenez R
  TITLE     Directed Evolution of a Bright Variant of mCherry: Suppression of 
            Nonradiative Decay by Fluorescence Lifetime Selections.
  JOURNAL   J Phys Chem B. 2022 Jun 16. doi: 10.1021/acs.jpcb.2c01956.
  PUBMED    35709514
REFERENCE   2  (bases 1 to 7591)
  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: "J Phys Chem
            B. 2022 Jun 16. doi: 10.1021/acs.jpcb.2c01956."
COMMENT     Sequence Label: piggyBAC-H2B mCherry-XL (R125K)
FEATURES             Location/Qualifiers
     source          1..7591
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..105)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(131..586)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(268..289)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     480..499
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     713..735
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     727..744
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     728..744
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   874..936
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        1440..1643
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1594..1614
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1640..1664
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1779..2156
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     CDS             2169..2879
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLKGTNFPSDGPVMQKKTMGSEASSERMYPEDGALK
                     GEVKYRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNRKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2314..2332)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2582..2601
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     promoter        2921..3132
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             3145..3413
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human 
                     T-cell leukemia virus (HTLV) type 1"
     CDS             3440..4039
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(3440..3459)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3936..3956
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(4099..4119)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    4161..4282
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4198..4217)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4252..4271
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   5045..5079
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(5570..5586)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5570..5586)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5583..5605)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5594..5610
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5618..5624,5625..5642,5643..5648))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5663..5684
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5801..5818)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5972..6560)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6052..6071)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6731..7522,7523..7591))
                     /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     7354..7373
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
       61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      121 agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
      181 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
      241 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
      301 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
      361 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
      421 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
      481 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
      541 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc
      601 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
      661 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
      721 cgacgttgta aaacgacggc cagtgagcgc gcctcgttca ttcacgtttt tgaacccgtg
      781 gaggacgggc agactcgcgg tgcaaatgtg ttttacagcg tgatggagca gatgaagatg
      841 ctcgacacgc tgcagaacac gcagctagat taaccctaga aagataatca tattgtgacg
      901 tacgttaaag ataatcatgt gtaaaattga cgcatgtgtt ttatcggtct gtatatcgag
      961 gtttatttat taatttgaat agatattaag ttttattata tttacactta catactaata
     1021 ataaattcaa caaacaattt atttatgttt atttatttat taaaaaaaac aaaaactcaa
     1081 aatttcttct ataaagtaac aaaactttta tgagggacag ccccccccca aagcccccag
     1141 ggatgtaatt acgtccctcc cccgctaggg ggcagcagcg agccgcccgg ggctccgctc
     1201 cggtccggcg ctccccccgc atccccgagc cggcagcgtg cggggacagc ccgggcacgg
     1261 ggaaggtggc acgggatcgc tttcctctga acgcttctcg ctgctctttg agcctgcaga
     1321 cacctggggg gatacgggga aaaggcctcc acggccacta gtattatgcc cagtacatga
     1381 ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgg
     1441 tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca cggggatttc
     1501 caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact
     1561 ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg cgtgtacggt
     1621 gggaggttta tataagcaga gctcgtttag tgaaccgtca gatcgcctgg agacgccatc
     1681 cacgctgttt tgacctccat agaagattct agagctagcg aattcgctac cggtctcgag
     1741 ctcaagcttc gaattctgca gtcgacggta ccgccaccat gccagagcca gcgaagtctg
     1801 ctcccgcccc gaaaaagggc tccaagaagg cggtgactaa ggcgcagaag aaaggcggca
     1861 agaagcgcaa gcgcagccgc aaggagagct attccatcta tgtgtacaag gttctgaagc
     1921 aggtccaccc tgacaccggc atttcgtcca aggccatggg catcatgaat tcgtttgtga
     1981 acgacatttt cgagcgcatc gcaggtgagg cttcccgcct ggcgcattac aacaagcgct
     2041 cgaccatcac ctccagggag atccagacgg ccgtgcgcct gctgctgcct ggggagttgg
     2101 ccaagcacgc cgtgtccgag ggtactaagg ccatcaccaa gtacaccagc gctaaggatc
     2161 ccaaaaaaat ggtgagcaag ggcgaggagg ataacatggc catcatcaag gagttcatgc
     2221 gcttcaaggt gcacatggag ggctccgtga acggccacga gttcgagatc gagggcgagg
     2281 gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggtggcc
     2341 ccctgccctt cgcctgggac atcctgtccc ctcagttcat gtacggctcc aaggcctacg
     2401 tgaagcaccc cgccgacatc cccgactact tgaagctgtc attccccgag ggcttcaagt
     2461 gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac cgtgacccag gactcctccc
     2521 tgcaggacgg cgagttcatc tacaaggtga agctgaaggg caccaacttt ccctccgacg
     2581 gccccgtaat gcagaagaag accatgggca gtgaggcctc ctccgagcgg atgtaccccg
     2641 aggacggcgc cctgaagggc gaggttaagt ataggctgaa gctgaaggac ggcggccact
     2701 acgacgctga ggtcaagacc acctacaagg ccaagaagcc cgtgcagctg cccggcgcct
     2761 acaacgtcaa ccgtaagttg gacatcacct cccacaacga ggactacacc atcgtggaac
     2821 agtacgaacg cgccgagggc cgccactcca ccggcggcat ggacgagctg tacaagtaag
     2881 cggccgcgaa ggatctgcga tcgctccggt gcccgtcagt gggcagagcg cacatcgccc
     2941 acagtccccg agaagttggg gggaggggtc ggcaattgaa cgggtgccta gagaaggtgg
     3001 cgcggggtaa actgggaaag tgatgtcgtg tactggctcc gcctttttcc cgagggtggg
     3061 ggagaaccgt atataagtgc agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc
     3121 gccagaacac agctgaagct tcgaggggct cgcatctctc cttcacgcgc ccgccgccct
     3181 acctgaggcc gccatccacg ccggttgagt cgcgttctgc cgcctcccgc ctgtggtgcc
     3241 tcctgaactg cgtccgccgt ctaggtaagt ttaaagctca ggtcgagacc gggcctttgt
     3301 ccggcgctcc cttggagcct acctagactc agccggctct ccacgctttg cctgaccctg
     3361 cttgctcaac tctacgtctt tgtttcgttt tctgttctgc gccgttacag atccaagctg
     3421 tgaccggcgc ctacgctaga tgaccgagta caagcccacg gtgcgcctcg ccacccgcga
     3481 cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg
     3541 ccacaccgtc gatccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct
     3601 cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc
     3661 ggtctggacc acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg
     3721 catggccgag ttgagcggtt cccggctggc cgcgcagcaa cagatggaag gcctcctggc
     3781 gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca
     3841 ccagggcaag ggtctgggca gcgccgtcgt gctccccgga gtggaggcgg ccgagcgcgc
     3901 cggggtgccc gccttcctgg agacctccgc gccccgcaac ctccccttct acgagcggct
     3961 cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac
     4021 ccgcaagccc ggtgcctgag tcgacaatca acctctggat tacaaaattt gtgaaagatt
     4081 gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc
     4141 tttgtatcat gcgttaacta aacttgttta ttgcagctta taatggttac aaataaagca
     4201 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     4261 ccaaactcat caatgtatct tatcatgtct ggaattgact caaatgatgt caattagtct
     4321 atcagaagct catctggtct cccttccggg ggacaagaca tccctgttta atatttaaac
     4381 agcagtgttc ccaaactggg ttcttatatc ccttgctctg gtcaaccagg ttgcagggtt
     4441 tcctgtcctc acaggaacga agtccctaaa gaaacagtgg cagccaggtt tagccccgga
     4501 attgactgga ttcctttttt agggcccatt ggtatggctt tttccccgta tccccccagg
     4561 tgtctgcagg ctcaaagagc agcgagaagc gttcagagga aagcgatccc gtgccacctt
     4621 ccccgtgccc gggctgtccc cgcacgctgc cggctcgggg atgcgggggg agcgccggac
     4681 cggagcggag ccccgggcgg ctcgctgctg ccccctagcg ggggagggac gtaattacat
     4741 ccctgggggc tttggggggg ggctgtccct gatatctata acaagaaaat atatatataa
     4801 taagttatca cgtaagtaga acatgaaata acaatataat tatcgtatga gttaaatctt
     4861 aaaagtcacg taaaagataa tcatgcgtca ttttgactca cgcggtcgtt atagttcaaa
     4921 atcagtgaca cttaccgcat tgacaagcac gcctcacggg agctccaagc ggcgactgag
     4981 atgtcctaaa tgcacagcga cggattcgcg ctatttagaa agagagagca atatttcaag
     5041 aatgcatgcg tcaattttac gcagactatc tttctagggt taatctagct gcatcaggat
     5101 catatcgtcg ggtctttttt ccggctcagt catcgcccaa gctggcgcta tctgggcatc
     5161 ggggaggaag aagcccgtgc cttttcccgc gaggttgaag cggcatggaa agagtttgcc
     5221 gaggatgact gctgctgcat tgacgttgag cgaaaacgca cgtttaccat gatgattcgg
     5281 gaaggtgtgg ccatgcacgc ctttaacggt gaactgttcg ttcaggccac ctgggatacc
     5341 agttcgtcgc ggcttttccg gacacagttc cggatggtca gcccgaagcg catcagcaac
     5401 ccgaacaata ccggcgacag ccggaactgc cgtgccggtg tgcagattaa tgacagcggt
     5461 gcggcgctgg gatattacgt cagcgaggac gggtatcctg gctggatgcc gcagaaatgg
     5521 acatggatac cccgtgagtt acccggcggg cgcgcttggc gtaatcatgg tcatagctgt
     5581 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     5641 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     5701 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
     5761 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     5821 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     5881 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     5941 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     6001 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     6061 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     6121 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     6181 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     6241 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     6301 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     6361 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     6421 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     6481 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     6541 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     6601 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     6661 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     6721 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     6781 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     6841 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     6901 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     6961 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     7021 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     7081 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     7141 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     7201 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     7261 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     7321 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     7381 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     7441 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     7501 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     7561 taagggcgac acggaaatgt tgaatactca t
//
