LOCUS       sequence-177081-        6752 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses TM-CIBN-BLITz1-TetR-VP16 of BLITz system.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6752)
  AUTHORS   Lee D, Creed M, Jung K, Stefanelli T, Wendler DJ, Oh WC, Mignocchi 
            NL, Luscher C, Kwon HB
  TITLE     Temporally precise labeling and control of neuromodulatory circuits 
            in the mammalian brain.
  JOURNAL   Nat Methods. 2017 May;14(5):495-503. doi: 10.1038/nmeth.4234. Epub 
            2017 Apr 3.
  PUBMED    28369042
REFERENCE   2  (bases 1 to 6752)
  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"; doi: 
            "10.1038/nmeth.4234"; journalName: "Nat Methods"; date: "2017-05"; 
            volume: "14"; issue: "5"; pages: "495-503"
COMMENT     Sequence Label: pCMV-TM-CIBN-BLITz1-TetR-VP16
FEATURES             Location/Qualifiers
     source          1..6752
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..978
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        1014..1032
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1014..1031
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     sig_peptide     1051..1110
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             1114..1143
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1147..1293
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     CDS             1318..1827
                     /codon_start=1
                     /product="N-terminal portion of the CIB1 transcription 
                     factor from Arabidopsis thaliana"
                     /label=CIBN
                     /note="binds to CRY2 (cryptochrome 2) that has absorbed 
                     blue light (Kennedy et al., 2010)"
                     /translation="MNGAIGGDLLLNFPDMSVLERQRAHLKYLNPTFDSPLAGFFADSS
                     MITGGEMDSYLSTAGLNLPMMYGETTVEGDSRLSISPETTLGTGNFKAAKFDTETKDCN
                     EAAKKMTMNRDDLVEEGEEEKSKITEQNNGSTKSIKKMKHKAKKEENNFSNDSSKVTKE
                     LEKTDYI"
     CDS             1903..1929
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     CDS             2725..2745
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             join(2746..3363,3364..3750)
                     /codon_start=1
                     /product="tetracycline-controlled transactivator, 
                     comprising a fusion of the tetracycline repressor TetR with
                     the C-terminal activation domain of herpes simplex virus 
                     VP16"
                     /label=tTA
                     /note="In the Tet-Off(R) system, tTA binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the absence of tetracycline or doxycycline."
                     /translation="SRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHVK
                     NKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTRP
                     TEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTTD
                     SMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGSAYSRARTKNNYGSTIE
                     GLLDLPDDDAPEEAGLAAPRLSFLPAGHTRRLSTAPPTDVSLGDELHLDGEDVAMAHAD
                     ALDDFDLDMLGDGDSPGPGFTPHDSAPYGALDMADFEFEQMFTDALGIDEYGG"
     primer_bind     complement(2816..2835)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(3759..3778)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3760..3778)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    3783..3917
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3807..3826)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3861..3880
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4030..4050)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4030..4048)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4069..4085)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4069..4085)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4082..4104)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4093..4109
                     /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(4117..4123,4124..4141,4142..4147))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4162..4183
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4300..4317)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4470..5058)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4550..4569)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5229..6020,6021..6089))
                     /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     5852..5871
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6090..6194)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6220..6675)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6357..6378)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6569..6588
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gccaattcaa tatggcgtat atggactcat gccaattcaa tatggtggat ctggacctgt
       61 gccaattcaa tatggcgtat atggactcgt gccaattcaa tatggtggat ctggacccca
      121 gccaattcaa tatggcggac ttggcaccat gccaattcaa tatggcggac ctggcactgt
      181 gccaactggg gaggggtcta cttggcacgg tgccaagttt gaggaggggt cttggccctg
      241 tgccaagtcc gccatattga attggcatgg tgccaataat ggcggccata ttggctatat
      301 gccaggatca atatataggc aatatccaat atggccctat gccaatatgg ctattggcca
      361 ggttcaatac tatgtattgg ccctatgcca tatagtattc catatatggg ttttcctatt
      421 gacgtagata gcccctccca atgggcggtc ccatatacca tatatggggc ttcctaatac
      481 cgcccatagc cactccccca ttgacgtcaa tggtctctat atatggtctt tcctattgac
      541 gtcatatggg cggtcctatt gacgtatatg gcgcctcccc cattgacgtc aattacggta
      601 aatggcccgc ctggctcaat gcccattgac gtcaatagga ccacccacca ttgacgtcaa
      661 tgggatggct cattgcccat tcatatccgt tctcacgccc cctattgacg tcaatgacgg
      721 taaatggccc acttggcagt acatcaatat ctattaatag taacttggca agtacattac
      781 tattggaagt acgccagggt acattggcag tactcccatt gacgtcaatg gcggtaaatg
      841 gcccgcgatg gctgccaagt acatccccat tgacgtcaat ggggaggggc aatgacgcaa
      901 atgggcgttc cattgacgta aatgggcggt aggcgtgcct aatgggaggt ctatataagc
      961 aatgctcgtt tagggaaccg ccattctgcc tggggacgtc ggagcaagct ttcatttagg
     1021 tgacactata gaatacaagc tacttgttct atggagacag acacactcct gctatgggta
     1081 ctgctgctct gggttccagg ttccactggt gacgaacaaa aactcatctc agaagaggat
     1141 ctgaatgctg tgggccagga cacgcaggag gtcatcgtgg tgccacactc cttgcccttt
     1201 aaggtggtgg tgatctcagc catcctggcc ctggtggtgc tcaccatcat ctcccttatc
     1261 atcctcatca tgctttggca gaagaagcca cgtggtggct ctggaggtct cgagggaatg
     1321 aatggagcta taggaggtga ccttttgctc aattttcctg acatgtcggt cctagagcgc
     1381 caaagggctc acctcaagta cctcaatccc acctttgatt ctcctctcgc cggcttcttt
     1441 gccgattctt caatgattac cggcggcgag atggacagct atctttcgac tgccggtttg
     1501 aatcttccga tgatgtacgg tgagacgacg gtggaaggtg attcaagact ctcaatttcg
     1561 ccggaaacga cgcttgggac tggaaatttc aaggcagcga agtttgatac agagactaag
     1621 gattgtaatg aggcggcgaa gaagatgacg atgaacagag atgacctagt agaagaagga
     1681 gaagaagaga agtcgaaaat aacagagcaa aacaatggga gcacaaaaag catcaagaag
     1741 atgaaacaca aagccaagaa agaagagaac aatttctcta atgattcatc taaagtgacg
     1801 aaggaattgg agaaaacgga ttatattcat gcatgcggtg gcggtggctc tggaggtggt
     1861 gggtccggag gaggcggccg ctccggtgga tccatgcttc aacttcctcc tcttgaacgt
     1921 cttactctcg agatgggcga gagcctgttc aagggaccta gggactacaa ccctatcagt
     1981 agcacaattt gtcacctgac caacgagagt gatggccaca caacaagcct gtacggcatc
     2041 gggttcggac cctttatcat caccaacaag cacctgttca ggcggaataa tggcactctg
     2101 ctggtgcaga gcctgcacgg ggtgttcaaa gtgaagaaca caaccactct gcagcagcac
     2161 ctgatcgatg ggcgggatat gatcatcatt aggatgccca aggacttccc cccttttcct
     2221 cagaaactga agttccgaga gccccagaga gaggagagaa tctgtctggt gaccacaaac
     2281 tttcaggagc tcggtagtgg tagtggggag tttctggcaa ccacactgga acggatcgag
     2341 aaaaatttcg tgattactga tccgagactg cctgacaacc caatcatttt tgcgagcgat
     2401 tccttcctgc agctgacaga atattctcgg gaagagatcc tggggcgcaa ttgccgtttt
     2461 ctgcagggac ccgagacaga ccgtgccact gttcggaaaa tcagagatgc tattgacaac
     2521 cagactgaag tgaccgttca gctgatcaat tataccaaga gcggcaagaa gttctggaac
     2581 gtgttccacc tgcagccgat gcgcgattat aagggcgacg tccagtactt cattggcgtg
     2641 cagctggatg gcaccgaacg tcttcatggc gccgctgagc gtgaggcggt ctgcctgatc
     2701 aaaaagacag cctttcagat tgctgagaac ctgtacttcc agggctctag attagataaa
     2761 agtaaagtga ttaacagcgc attagagctg cttaatgagg tcggaatcga aggtttaaca
     2821 acccgtaaac tcgcccagaa gctaggtgta gagcagccta cattgtattg gcatgtaaaa
     2881 aataagcggg ctttgctcga cgccttagcc attgagatgt tagataggca ccatactcac
     2941 ttttgccctt tagaagggga aagctggcaa gattttttac gtaataacgc taaaagtttt
     3001 agatgtgctt tactaagtca tcgcgatgga gcaaaagtac atttaggtac acggcctaca
     3061 gaaaaacagt atgaaactct cgaaaatcaa ttagcctttt tatgccaaca aggtttttca
     3121 ctagagaatg cattatatgc actcagcgct gtggggcatt ttactttagg ttgcgtattg
     3181 gaagatcaag agcatcaagt cgctaaagaa gaaagggaaa cacctactac tgatagtatg
     3241 ccgccattat tacgacaagc tatcgaatta tttgatcacc aaggtgcaga gccagccttc
     3301 ttattcggcc ttgaattgat catatgcgga ttagaaaaac aacttaaatg tgaaagtggg
     3361 tccgcgtaca gccgcgcgcg tacgaaaaac aattacgggt ctaccatcga gggcctgctc
     3421 gatctcccgg acgacgacgc ccccgaagag gcggggctgg cggctccgcg cctgtccttt
     3481 ctccccgcgg gacacacgcg cagactgtcg acggcccccc cgaccgatgt cagcctgggg
     3541 gacgagctcc acttagacgg cgaggacgtg gcgatggcgc atgccgacgc gctagacgat
     3601 ttcgatctgg acatgttggg ggacggggat tccccgggtc cgggatttac cccccacgac
     3661 tccgccccct acggcgctct ggatatggcc gacttcgagt ttgagcagat gtttaccgat
     3721 gcccttggaa ttgacgagta cggtgggtag gctagcctcc ctatagtgag tcgtattacg
     3781 tagatccaga catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt
     3841 gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa
     3901 gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg
     3961 aggtgtggga ggttttttaa ttcgcgtccg cggcgccaat gcattgggcc cggtacccag
     4021 cttttgttcc ctttagtgag ggttaattgc gcgcttggcg taatcatggt catagctgtt
     4081 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg ggagcataaa
     4141 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     4201 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     4261 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     4321 ctcggtcgtt cggctgcggg agcggtatca gctcactcaa aggcggtaat acggttatcc
     4381 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     4441 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     4501 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     4561 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     4621 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     4681 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     4741 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     4801 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     4861 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     4921 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     4981 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     5041 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     5101 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     5161 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     5221 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     5281 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     5341 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     5401 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     5461 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     5521 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     5581 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     5641 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     5701 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     5761 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     5821 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     5881 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     5941 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     6001 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     6061 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     6121 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     6181 ataggggttc cgcgcacatt tccccgaaaa gtgccaccta aattgtaagc gttaatattt
     6241 tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa
     6301 tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag
     6361 tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg
     6421 tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga
     6481 ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg
     6541 gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg
     6601 cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc
     6661 cgctacaggg cgcgtcccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg
     6721 tgcgggcctc ttcgctatta cgccagtcga cc
//
