LOCUS       sequence-434071-        6545 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  NLS-Gal4(1-65)-iLight2-VP16-T2A-mTagBFP2 construct expression in 
            mammalian cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6545)
  AUTHORS   Baloban M, Kasatkina LA, Verkhusha VV
  TITLE     iLight2: A near-infrared optogenetic tool for gene transcription 
            with low background activation.
  JOURNAL   Protein Sci. 2024 May;33(5):e4993. doi: 10.1002/pro.4993.
  PUBMED    38647395
REFERENCE   2  (bases 1 to 6545)
  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: "Protein 
            Sci."; date: "2024-05"; volume: "33(5):e4993. doi"; pages: " 
            10.1002/pro.4993"
COMMENT     Sequence Label: pCMVd1-NLS-GAL4-iLight2-VP16-T2A-mTagBFP2
FEATURES             Location/Qualifiers
     source          1..6545
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    104..225
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(141..160)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     195..214
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(232..687)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(369..390)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     581..600
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        714..818
                     /gene="bla"
                     /label=AmpR promoter
     promoter        820..1177
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        820..1011
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(843..863)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1028..1163
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             1212..2006
                     /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             1212..2006
                     /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(1266..1285)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1876..1895
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(2194..2213)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2238..2285
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      2614..3202
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3103..3122
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3273..3372
                     /label=CMVd1 promoter
                     /note="truncated human cytomegalovirus (CMV) immediate 
                     early promoter that yields slightly reduced expression 
                     relative to the full-length promoter"
     primer_bind     3323..3343
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             5504..5737
                     /codon_start=1
                     /gene="UL48"
                     /product="transcriptional activation domain of herpes 
                     simplex virus protein VP16 (Triezenberg et al., 1988; 
                     Cousens et al., 1989)"
                     /label=VP16 AD
                     /translation="APPTDVSLGDELHLDGEDVAMAHADALDDFDLDMLGDGDSPGPGF
                     TPHDSAPYGALDMADFEFEQMFTDALGIDEYGG"
     CDS             5756..5809
                     /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             5816..6514
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="SELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKVV
                     EGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTAT
                     QDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALKL
                     VGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYCD
                     LPSKLGHKLN"
ORIGIN
        1 cataatcagc cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct
       61 ccccctgaac ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc
      121 ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc
      181 actgcattct agttgtggtt tgtccaaact catcaatgta tcttaaggcg taaattgtaa
      241 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
      301 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
      361 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
      421 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
      481 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
      541 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
      601 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
      661 cgcttaatgc gccgctacag ggcgcgtcag gtggcacttt tcggggaaat gtgcgcggaa
      721 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
      781 cctgataaat gcttcaataa tattgaaaaa ggaagagtcc tgaggcggaa agaaccagct
      841 gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
      901 gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc
      961 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac
     1021 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     1081 aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta
     1141 gtgaggaggc ttttttggag gcctaggctt ttgcaaagat cgatcaagag acaggatgag
     1201 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     1261 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     1321 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     1381 tgaatgaact gcaagacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     1441 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     1501 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     1561 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     1621 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     1681 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcga
     1741 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     1801 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     1861 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     1921 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     1981 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     2041 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     2101 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     2161 ggagttcttc gcccacccta gggggaggct aactgaaaca cggaaggaga caataccgga
     2221 aggaacccgc gctatgacgg caataaaaag acagaataaa acgcacggtg ttgggtcgtt
     2281 tgttcataaa cgcggggttc ggtcccaggg ctggcactct gtcgataccc caccgagacc
     2341 ccattggggc caatacgccc gcgtttcttc cttttcccca ccccaccccc caagttcggg
     2401 tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag cctcaggtta
     2461 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     2521 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     2581 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     2641 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     2701 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     2761 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     2821 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     2881 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     2941 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     3001 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     3061 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     3121 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     3181 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     3241 ttgctggcct tttgctcaca tgttctttcc tgccaaaatc aacgggactt tccaaaatgt
     3301 cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat
     3361 ataagcagag ctggtttagt gaaccgtcag atccgctagc atggcgcgca agaagaagag
     3421 gaagtccgga ggtggtggaa agctactgtc ttctatcgaa caagcatgcg atatttgccg
     3481 acttaaaaag ctcaagtgct ccaaagaaaa accgaagtgc gccaagtgtc tgaagaacaa
     3541 ctgggagtgt cgctactctc ccaaaaccaa aaggtctccg ctgactaggg cacatctgac
     3601 agaagtggaa tcaaggctag aaagactgga acagactagt gctggtgggg aattcgccgc
     3661 cgacctgggc tctgacgata tcagcaagct gatcgccgcc tgcgatcagg agccaatcca
     3721 catccccaat gccatccagc catttggcgc catgctgatc gtggagaagg acacacagca
     3781 gatcgtgtac gcctctgcca acagcgccga gtacttcagc gtggccgaca ataccatcca
     3841 cgagctgtcc gatttcaagc aggccaacat caattctctg ctgcccgagc agctgatcag
     3901 cggcctgaca tccgccatct ctgagaacga gcctatctgg gtggagaccg acaggctgag
     3961 ctttctgggc tggcgccacg agaactacta tatcatcgag gtggagagat accacgtgca
     4021 gacatccaat tggttcgaga tccagtttca gcgggccttc cagaagctga gaaactgtaa
     4081 gacccacaac gatctgatca ataccctgac acggctgatc caggagatca gcggctacga
     4141 cagagtgatg atctatcagt tcgatcccga gtggaatggc agagtgatcg ccgagagcgt
     4201 gagacagctg tttacctcca tgctgaacca ccacttccca gcctctgaca tccctgcaca
     4261 ggccagggcc atgtacagca tcaacccaat ccgcatcatc cccgatgtga atgccgagcc
     4321 ccagcctctg cacatgatcc acaagccaca gaacacagag gccgtgaatc tgtcctgcgg
     4381 cgtgctgagg gccgtgtctc cactgcacat gcagtatctg cgcaactttg gcgtgtctgc
     4441 cagcacctcc atcggcatct tcaatgagga caagctgtgg ggcatcgtgg cctgtcacca
     4501 cacaaagcct agggccatcg gccggagaat caggcacctg ctggtgcgca ccgtggagtt
     4561 tgcagcagag cgcctgtggc tgatccactc caggaatgtg gagcggtaca tggtgacagt
     4621 gcaggcagcc cgggagcagc tgtctaccac agccgacgat aagcacagct cccacgagat
     4681 cgtgatcgag cacgccgccg actggtgcaa gctgttccgg tgtgatggcg tgggctacct
     4741 gagaggcgag gagctgacca catatggcga gacccctgat cagaccacaa tcaacaagct
     4801 ggtggagtgg ctggaggaga atggcaagaa gagcctgttt tggcactccc acatgctgaa
     4861 ggaggacgca cctggactgc tgccagatgg cagccggttc gcaggactgc tggccatccc
     4921 actgaagtct gacgccgatc tgtttagcta cctgctgctg ttcagggtgg cacagaacga
     4981 ggtgcgcaca tgggcaggca agcctgagaa gctgtccgtg gagacctcta caggcaccat
     5041 ggtgggccca cggaagtctt ttgaggcctg gcaggacgag gtgagcggca agtcccagcc
     5101 ttggagaacc gcacagctgt atgcagcccg ggacatcgcc cgggacctgc tgatcgtggc
     5161 cgatagcatg cagctgaacc tgctgaatga ccagctggcc gatgccaacg agaatctgga
     5221 gaagctggcc tccttcgacg atctgacctc tggcggcggt accagcgggg gtggtggatc
     5281 aggtggagga ggttctggag gtggtggatc aggaggaggt ggttctggag gtggtggatc
     5341 aggaggaaga tctgcgtaca gccgcgcgcg tacgaaaaac aattacgggt ctaccatcga
     5401 gggcctgctc gatctcccgg acgacgacgc ccccgaagag gcggggctgg cggctccgcg
     5461 cctgtccttt ctccccgcgg gacacacgcg cagactgtcg acggcccccc cgaccgatgt
     5521 cagcctgggg gacgagctcc acttagacgg cgaggacgtg gcgatggcgc atgccgacgc
     5581 gctagacgat ttcgatctgg acatgttggg ggacggggat tcccctggtc cgggatttac
     5641 cccccacgac tccgccccct acggcgctct ggatatggcc gacttcgagt ttgagcagat
     5701 gtttaccgat gcccttggaa ttgacgagta cggtggggct cgagataggc gcgccgaggg
     5761 cagaggaagt ctgctaacat gcggtgacgt cgaggagaat cctggcccaa ctagtagcga
     5821 gctgattaag gagaacatgc acatgaagct gtacatggag ggcaccgtgg acaaccatca
     5881 cttcaagtgc acatccgagg gcgaaggcaa gccctacgag ggcacccaga ccatgagaat
     5941 caaggtggtc gagggcggcc ctctcccctt cgccttcgac atcctggcta ctagcttcct
     6001 ctacggcagc aagaccttca tcaaccacac ccagggcatc cccgacttct tcaagcagtc
     6061 cttccctgag ggcttcacat gggagagagt caccacatac gaagacgggg gcgtgctgac
     6121 cgctacccag gacaccagcc tccaggacgg ctgcctcatc tacaacgtca agatcagagg
     6181 ggtgaacttc acatccaacg gccctgtgat gcagaagaaa acactcggct gggaggcctt
     6241 caccgagacg ctgtaccccg ctgacggcgg cctggaaggc agaaacgaca tggccctgaa
     6301 gctcgtgggc gggagccatc tgatcgcaaa cgccaagacc acatatagat ccaagaaacc
     6361 cgctaagaac ctcaagatgc ctggcgtcta ctatgtggac tacagactgg aaagaatcaa
     6421 ggaggccaac aacgagacct acgtcgagca gcacgaggtg gcagtggcca gatactgcga
     6481 cctccctagc aaactggggc acaagcttaa ttaacccggg atccaccggg gccgcgactc
     6541 tagat
//
