LOCUS       sequence-417724-        5221 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  NIR optogenetic module for conditional protein splicing of split 
            mCherry (C-part).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5221)
  AUTHORS   Karasev MM, Verkhusha VV, Shcherbakova DM
  TITLE     Near-Infrared Optogenetic Module for Conditional Protein Splicing.
  JOURNAL   J Mol Biol. 2023 Dec 15;435(24):168360. doi: 
            10.1016/j.jmb.2023.168360. Epub 2023 Nov 8.
  PUBMED    37949312
REFERENCE   2  (bases 1 to 5221)
  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 Mol 
            Biol."; date: "2023-12-15"; volume: "435(24):168360. doi"; pages: " 
            10.1016/j.jmb.2023.168360. Epub 2023 Nov 8"
COMMENT     Sequence Label: pN1-CMV-Aff6_V18FdN-IntC-mCherryC-T2A-EGFP (C2)
FEATURES             Location/Qualifiers
     source          1..5221
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     159..178
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(477..496)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    521..568
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      897..1485
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1386..1405
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        1660..1963
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1964..2167
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2118..2138
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             2712..2765
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             join(2766..2768,2769..2771,2772..3485)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2811..2832)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3072..3091)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3419..3440
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3608..3729
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3645..3664)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3699..3718
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3736..4191)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3873..3894)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4085..4104
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4218..4322
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4324..4681
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4324..4515
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4347..4367)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4532..4667
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             join(4716..5221,1..289)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             join(4716..5221,1..289)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4770..4789)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 actgttcgcc aggctcaagg cgagcatgcc cgacggcgag gatctcgtcg tgacccatgg
       61 cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg
      121 tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc
      181 tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc
      241 cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg
      301 gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc
      361 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
      421 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ctagggggag gctaactgaa
      481 acacggaagg agacaatacc ggaaggaacc cgcgctatga cggcaataaa aagacagaat
      541 aaaacgcacg gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca gggctggcac
      601 tctgtcgata ccccaccgag accccattgg ggccaatacg cccgcgtttc ttccttttcc
      661 ccaccccacc ccccaagttc gggtgaaggc ccagggctcg cagccaacgt cggggcggca
      721 ggccctgcca tagcctcagg ttactcatat atactttaga ttgatttaaa acttcatttt
      781 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
      841 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
      901 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
      961 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     1021 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     1081 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     1141 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     1201 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     1261 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     1321 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     1381 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     1441 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     1501 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     1561 tcccctgatt ctgtggataa ccgtattacc gccatgcatt agttattaat agtaatcaat
     1621 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
     1681 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
     1741 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
     1801 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
     1861 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
     1921 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca
     1981 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
     2041 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
     2101 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
     2161 cagagctggt ttagtgaacc gtcagatccg ctagtagatc tagcaccatg aagtttaaca
     2221 aggagaagtg gcaggctatt aaggagattg ccttcctgcc taacctgaac cagccccagc
     2281 acctggcttt tattgtgtct ctgctggacg acccttctca gtctgctaac ctgctggctg
     2341 aggctaagaa gctgaacgac gctcaggctc ctaagggatc cgaactgatt gatattgaag
     2401 ttagcggtaa ccacctgttc tatgccaatg atattctgac ccataatagc tcgagccccg
     2461 aggacggcgc cctgaagggc gagatcaagc agaggctgaa gctgaaggac ggcggccact
     2521 acgacgctga ggtcaagacc acctacaagg ccaagaagcc cgtgcagctg cccggcgcct
     2581 acaacgtcaa catcaagttg gacatcacct cccacaacga ggactacacc atcgtggaac
     2641 agtacgaacg cgccgagggc cgccactcca ccggcggcat ggacgagctg tacaaggaat
     2701 tcggtagcgg cgagggcaga ggaagtctgc taacatgcgg tgatgtcgag gagaatcctg
     2761 gcccaatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     2821 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     2881 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     2941 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     3001 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     3061 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     3121 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     3181 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     3241 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     3301 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     3361 accactacct gagcacccag tccaagctga gcaaagaccc caacgagaag cgcgatcaca
     3421 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     3481 agtaaagcgg ccgcgactct agatcataat cagccatacc acatttgtag aggttttact
     3541 tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt
     3601 tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     3661 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     3721 tgtatcttaa ggcgtaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     3781 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     3841 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     3901 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     3961 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     4021 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     4081 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     4141 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca
     4201 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     4261 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     4321 gtcctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     4381 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     4441 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     4501 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     4561 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     4621 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     4681 agatcgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg
     4741 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     4801 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     4861 tcaagaccga cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt
     4921 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     4981 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     5041 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     5101 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     5161 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     5221 a
//
