LOCUS       sequence-364141-        6974 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Optogenetic PHR domain coupled to GFP and p65 activation domain; 
            binds to CIBN upon blue light exposure.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6974)
  AUTHORS   Trojanowski J, Frank L, Rademacher A, Mucke N, Grigaitis P, Rippe K
  TITLE     Transcription activation is enhanced by multivalent interactions 
            independent of phase separation.
  JOURNAL   Mol Cell. 2022 May 4. pii: S1097-2765(22)00328-8. doi: 
            10.1016/j.molcel.2022.04.017.
  PUBMED    35537448
REFERENCE   2  (bases 1 to 6974)
  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: "Mol Cell. 
            2022 May 4. pii: S1097-2765(22)00328-8. doi: 
            10.1016/j.molcel.2022.04.017."
COMMENT     Sequence Label: PHR-GFP-p65
FEATURES             Location/Qualifiers
     source          1..6974
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             625..2118
                     /codon_start=1
                     /product="N-terminal photolyase homology region of 
                     Arabidopsis thaliana cryptochrome 2"
                     /label=CRY2PHR
                     /note="binds to CIB1 upon exposure to blue light (Kennedy 
                     et al., 2010)"
                     /translation="MKMDKKTIVWFRRDLRIEDNPALAAAAHEGSVFPVFIWCPEEEGQ
                     FYPGRASRWWMKQSLAHLSQSLKALGSDLTLIKTHNTISAILDCIRVTGATKVVFNHLY
                     DPVSLVRDHTVKEKLVERGISVQSYNGDLLYEPWEIYCEKGKPFTSFNSYWKKCLDMSI
                     ESVMLPPPWRLMPITAAAEAIWACSIEELGLENEAEKPSNALLTRAWSPGWSNADKLLN
                     EFIEKQLIDYAKNSKKVVGNSTSLLSPYLHFGEISVRHVFQCARMKQIIWARDKNSEGE
                     ESADLFLRGIGLREYSRYICFNFPFTHEQSLLSHLRFFPWDADVDKFKAWRQGRTGYPL
                     VDAGMRELWATGWMHNRIRVIVSSFAVKFLLLPWKWGMKYFWDTLLDADLECDILGWQY
                     ISGSIPDGHELDRLDNPALQGAKYDPEGEYIRQWLPELARLPTEWIHHPWDAPLTVLKA
                     SGVELGTNYAKPIVDIDTARELLAKAISRTREAQIMIGAA"
     CDS             join(2128..2130,2131..2133,2134..2844)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2128..2130,2131..2133,2134..2844)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2173..2194)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2434..2453)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2781..2802
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2878..2898
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3388..3744
                     /codon_start=1
                     /product="transcriptional activation domain of human RelA, 
                     also known as p65 (O'Shea and Perkins, 2008)"
                     /label=RelA (p65) AD
                     /translation="PTQAGEGTLSEALLQLQFDDEDLGALLGNSTDPAVFTDLASVDNS
                     EFQQLLNQGIPVAPHTTEPMLMEYPEAITRLVTGAQRPPDPAPAPLGAPGLPNGLLSGD
                     EDFSSIADMDFSALL"
     polyA_signal    3762..3883
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3799..3818)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3853..3872
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3890..4345)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4027..4048)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4239..4258
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4372..4476
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4478..4835
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4478..4669
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4501..4521)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4686..4821
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4870..5664
                     /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             4870..5664
                     /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(4924..4943)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5534..5553
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5852..5871)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5896..5943
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6272..6860
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6761..6780
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgaggc caccatgaag atggacaaaa agactatagt ttggtttaga
      661 agagacctaa ggattgagga taatcctgca ttagcagcag ctgctcacga aggatctgtt
      721 tttcctgtct tcatttggtg tcctgaagaa gaaggacagt tttatcctgg aagagcttca
      781 agatggtgga tgaaacaatc acttgctcac ttatctcaat ccttgaaggc tcttggatct
      841 gacctcactt taatcaaaac ccacaacacg atttcagcga tcttggattg tatccgcgtt
      901 accggtgcta caaaagtcgt ctttaaccac ctctatgatc ctgtttcgtt agttcgggac
      961 cataccgtaa aggagaagct ggtggaacgt gggatctctg tgcaaagcta caatggagat
     1021 ctattgtatg aaccgtggga gatatactgc gaaaagggca aaccttttac gagtttcaat
     1081 tcttactgga agaaatgctt agatatgtcg attgaatccg ttatgcttcc tcctccttgg
     1141 cggttgatgc caataactgc agcggctgaa gcgatttggg cgtgttcgat tgaagaacta
     1201 gggctggaga atgaggccga gaaaccgagc aatgcgttgt taactagagc ttggtctcca
     1261 ggatggagca atgctgataa gttactaaat gagttcatcg agaagcagtt gatagattat
     1321 gcaaagaaca gcaagaaagt tgttgggaat tctacttcac tactttctcc gtatctccat
     1381 ttcggggaaa taagcgtcag acacgttttc cagtgtgccc ggatgaaaca aattatatgg
     1441 gcaagagata agaacagtga aggagaagaa agtgcagatc tttttcttag gggaatcggt
     1501 ttaagagagt attctcggta tatatgtttc aacttcccgt ttactcacga gcaatcgttg
     1561 ttgagtcatc ttcggttttt cccttgggat gctgatgttg ataagttcaa ggcctggaga
     1621 caaggcagga ccggttatcc gttggtggat gccggaatga gagagctttg ggctaccgga
     1681 tggatgcata acagaataag agtgattgtt tcaagctttg ctgtgaagtt tcttctcctt
     1741 ccatggaaat ggggaatgaa gtatttctgg gatacacttt tggatgctga tttggaatgt
     1801 gacatccttg gctggcagta tatctctggg agtatccccg atggccacga gcttgatcgc
     1861 ttggacaatc ccgcgttaca aggcgccaaa tatgacccag aaggtgagta cataaggcaa
     1921 tggcttcccg agcttgcgag attgccaact gaatggatcc atcatccatg ggacgctcct
     1981 ttaaccgtac tcaaagcttc tggtgtggaa ctcggaacaa actatgcgaa acccattgta
     2041 gacatcgaca cagctcgtga gctactagct aaagctattt caagaacccg tgaagcacag
     2101 atcatgatcg gagcagcagc ccgggtcatg gtgagcaagg gcgaggagct gttcaccggg
     2161 gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc
     2221 ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc
     2281 ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc
     2341 ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa
     2401 ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc
     2461 gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc
     2521 aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc
     2581 tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac
     2641 atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac
     2701 ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac
     2761 cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact
     2821 ctcggcatgg acgagctgta caagagcggc cgcagggggc gtcgactggg tacccctcct
     2881 aagaagaaga ggaaggttgc ccccaccatg gataaaaaga agagaaaggc ccccccgacc
     2941 gatgtcagcc tgggggacga gctccagtac ctgcccgaca ccgacgaccg gcaccggatc
     3001 gaggaaaagc ggaagcggac ctacgagaca ttcaagagca tcatgaagaa gtcccccttc
     3061 agcggcccca ccgaccctag acctccacct agaagaatcg ccgtgcccag cagatccagc
     3121 gccagcgtgc caaaacctgc cccccagcct taccccttca ccagcagcct gagcaccatc
     3181 aactacgacg agttccctac catggtgttc cccagcggcc agatctctca ggcctctgct
     3241 ctggctccag cccctcctca ggtgctgcct caggctcctg ctcctgcacc agctccagcc
     3301 atggtgtctg cactggctca ggcaccagca cccgtgcctg tgctggctcc tggacctcca
     3361 caggctgtgg ctccaccagc ccctaaacct acacaggccg gcgagggcac actgtctgaa
     3421 gctctgctgc agctgcagtt cgacgacgag gatctgggag ccctgctggg aaacagcacc
     3481 gatcctgccg tgttcaccga cctggccagc gtggacaaca gcgagttcca gcagctgctg
     3541 aaccagggca tccctgtggc ccctcacacc accgagccca tgctgatgga ataccccgag
     3601 gccatcaccc ggctcgtgac aggcgctcag aggcctcctg atccagctcc tgcccctctg
     3661 ggagcaccag gcctgcctaa tggactgctg tctggcgacg aggacttcag ctctatcgcc
     3721 gatatggatt tctcagcctt gctgtaacaa ttgttgttgt taacttgttt attgcagctt
     3781 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     3841 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaaggcgta aattgtaagc
     3901 gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa
     3961 taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt
     4021 gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg
     4081 cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt
     4141 ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga
     4201 gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg
     4261 ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg
     4321 cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
     4381 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     4441 tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag aaccagctgt
     4501 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     4561 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     4621 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     4681 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     4741 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     4801 gaggaggctt ttttggaggc ctaggctttt gcaaagatcg atcaagagac aggatgagga
     4861 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     4921 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     4981 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     5041 aatgaactgc aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     5101 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     5161 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     5221 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     5281 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     5341 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgagc
     5401 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     5461 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     5521 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     5581 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     5641 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     5701 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     5761 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     5821 agttcttcgc ccaccctagg gggaggctaa ctgaaacacg gaaggagaca ataccggaag
     5881 gaacccgcgc tatgacggca ataaaaagac agaataaaac gcacggtgtt gggtcgtttg
     5941 ttcataaacg cggggttcgg tcccagggct ggcactctgt cgatacccca ccgagacccc
     6001 attggggcca atacgcccgc gtttcttcct tttccccacc ccacccccca agttcgggtg
     6061 aaggcccagg gctcgcagcc aacgtcgggg cggcaggccc tgccatagcc tcaggttact
     6121 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     6181 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     6241 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     6301 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     6361 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     6421 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     6481 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     6541 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     6601 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     6661 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     6721 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     6781 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     6841 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     6901 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     6961 ttaccgccat gcat
//
