LOCUS       sequence-177689-        6211 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Reporter construct.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6211)
  AUTHORS   Wang W, Wildes CP, Pattarabanjird T, Sanchez MI, Glober GF, Matthews
            GA, Tye KM, Ting AY
  TITLE     A light- and calcium-gated transcription factor for imaging and 
            manipulating activated neurons.
  JOURNAL   Nat Biotechnol. 2017 Jun 26. doi: 10.1038/nbt.3909.
  PUBMED    28650461
REFERENCE   2  (bases 1 to 6211)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Biotechnol. 2017 Jun 26. doi: 10.1038/nbt.3909."
COMMENT     Sequence Label: TRE-ChrimsonR-mCherry
FEATURES             Location/Qualifiers
     source          1..6211
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     protein_bind    176..194
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    211..229
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    247..265
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    283..301
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    318..336
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    354..372
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     promoter        384..422
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     419..443
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      471..480
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             477..1526
                     /codon_start=1
                     /product="red light-drivable channelrhodopsin from 
                     Chlamydomonas noctigama (Klapoetke et al., 2014)"
                     /label=ChrimsonR
                     /note="K176R mutant with accelerated closing kinetics"
                     /translation="MAELISSATRSLFAAGGINPWPNPYHHEDMGCGGMTPTGECFSTE
                     WWCDPSYGLSDAGYGYCFVEATGGYLVVGVEKKQAWLHSRGTPGEKIGAQVCQWIAFSI
                     AIALLTFYGFSAWKATCGWEEVYVCCVEVLFVTLEIFKEFSSPATVYLSTGNHAYCLRY
                     FEWLLSCPVILIRLSNLSGLKNDYSKRTMGLIVSCVGMIVFGMAAGLATDWLKWLLYIV
                     SCIYGGYMYFQAAKCYVEANHSVPKGHCRMVVKLMAYAYFASWGSYPILWAVGPEGLLK
                     LSPYANSIGHSICDIIAKEFWTFLAHHLRIKIHEHILIHGDIRKTTKMEIGGEEVEVEE
                     FVEEEDEDTV"
     regulatory      1590..1599
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1596..2306
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(1741..1759)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2009..2028
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    2338..2926
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2391..2411)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2809..2820)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2958..3434
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3157..3176)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3474..3614
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3474..3603
                     /label=AAV2 ITR
     rep_origin      3689..4144
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3776..3795)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3986..4007
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4161..4180)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4280..4302
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4340..4358)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4426..4530
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4531..4599,4600..5391)
                     /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     complement(4749..4768)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5562..6150
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6051..6070
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtccttca ggtccgaggt tctagacgag tttactccct
      181 atcagtgata gagaacgatg tcgagtttac tccctatcag tgatagagaa cgtatgtcga
      241 gtttactccc tatcagtgat agagaacgta tgtcgagttt actccctatc agtgatagag
      301 aacgtatgtc gagtttatcc ctatcagtga tagagaacgt atgtcgagtt tactccctat
      361 cagtgataga gaacgtatgt cgaggtaggc gtgtacggtg ggaggcctat ataagcagag
      421 ctcgtttagt gaaccgtcag atcgcaaagg gcgaattcga tccaccggtc gccaccatgg
      481 ctgagctgat cagcagcgcc accagatctc tgtttgccgc cggaggcatc aacccttggc
      541 ctaaccccta ccaccacgag gacatgggct gtggaggaat gacacctaca ggcgagtgct
      601 tcagcaccga gtggtggtgt gacccttctt acggactgag cgacgccgga tacggatatt
      661 gcttcgtgga ggccacaggc ggctacctgg tcgtgggagt ggagaagaag caggcttggc
      721 tgcacagcag aggcacacca ggagaaaaga tcggcgccca ggtctgccag tggattgctt
      781 tcagcatcgc catcgccctg ctgacattct acggcttcag cgcctggaag gccacttgcg
      841 gttgggagga ggtctacgtc tgttgcgtcg aggtgctgtt cgtgaccctg gagatcttca
      901 aggagttcag cagccccgcc acagtgtacc tgtctaccgg caaccacgcc tattgcctgc
      961 gctacttcga gtggctgctg tcttgccccg tgatcctgat cagactgagc aacctgagcg
     1021 gcctgaagaa cgactacagc aagcggacca tgggcctgat cgtgtcttgc gtgggaatga
     1081 tcgtgttcgg catggccgca ggactggcta ccgattggct caagtggctg ctgtatatcg
     1141 tgtcttgcat ctacggcggc tacatgtact tccaggccgc caagtgctac gtggaagcca
     1201 accacagcgt gcctaaaggc cattgccgca tggtcgtgaa gctgatggcc tacgcttact
     1261 tcgcctcttg gggcagctac ccaatcctct gggcagtggg accagaagga ctgctgaagc
     1321 tgagccctta cgccaacagc atcggccaca gcatctgcga catcatcgcc aaggagtttt
     1381 ggaccttcct ggcccaccac ctgaggatca agatccacga gcacatcctg atccacggcg
     1441 acatccggaa gaccaccaag atggagatcg gaggcgagga ggtggaagtg gaagagttcg
     1501 tggaggagga ggacgaggac acagtggcgg caccggtagt agcagtgagc ggtggaagcg
     1561 gttctggaag tgctcgggat ccaccggtcg ccaccatggt gagcaagggc gaggaggata
     1621 acatggccat catcaaggag ttcatgcgct tcaaggtgca catggagggc tccgtgaacg
     1681 gccacgagtt cgagatcgag ggcgagggcg agggccgccc ctacgagggc acccagaccg
     1741 ccaagctgaa ggtgaccaag ggtggccccc tgcccttcgc ctgggacatc ctgtcccctc
     1801 agttcatgta cggctccaag gcctacgtga agcaccccgc cgacatcccc gactacttga
     1861 agctgtcctt ccccgaggga ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg
     1921 tggtgaccgt gacccaggac tcctccctgc aggacggcga gttcatctac aaggtgaagc
     1981 tgcgcggcac caacttcccc tccgacggcc ccgtaatgca gaagaagacc atgggctggg
     2041 aggcctcctc cgagcggatg taccccgagg acggcgccct gaagggcgag atcaagcaga
     2101 ggctgaagct gaaggacggc ggccactacg acgctgaggt caagaccacc tacaaggcca
     2161 agaagcccgt gcagctgccc ggcgcctaca acgtcaacat caagttggac atcacctccc
     2221 acaacgagga ctacaccatc gtggaacagt acgaacgcgc cgagggccgc cactccaccg
     2281 gcggcatgga cgagctgtac aagtaaacta gtaccggtag atatcaagct tatcgataat
     2341 caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct
     2401 tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg
     2461 gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg
     2521 cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt
     2581 tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt
     2641 gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg
     2701 ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc
     2761 tatgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat
     2821 ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc
     2881 cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcatcg ataccgagcg
     2941 ctgctcgaga gatctacggg tggcatccct gtgacccctc cccagtgcct ctcctggccc
     3001 tggaagttgc cactccagtg cccaccagcc ttgtcctaat aaaattaagt tgcatcattt
     3061 tgtctgacta ggtgtccttc tataatatta tggggtggag gggggtggta tggagcaagg
     3121 ggcaagttgg gaagacaacc tgtagggcct gcggggtcta ttgggaacca agctggagtg
     3181 cagtggcaca atcttggctc actgcaatct ccgcctcctg ggttcaagcg attctcctgc
     3241 ctcagcctcc cgagttgttg ggattccagg catgcatgac caggctcagc taatttttgt
     3301 ttttttggta gagacggggt ttcaccatat tggccaggct ggtctccaac tcctaatctc
     3361 aggtgatcta cccaccttgg cctcccaaat tgctgggatt acaggcgtga accactgctc
     3421 ccttccctgt ccttctgatt ttgtaggtaa ccacgtgcgg accgagcggc cgcaggaacc
     3481 cctagtgatg gagttggcca ctccctctct gcgcgctcgc tcgctcactg aggccgggcg
     3541 accaaaggtc gcccgacgcc cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg
     3601 cagctgcctg caggggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc
     3661 acaccgcata cgtcaaagca accatagtac gcgccctgta gcggcgcatt aagcgcggcg
     3721 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     3781 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     3841 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     3901 gatttgggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     3961 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     4021 cctatctcgg gctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     4081 aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgtttaca
     4141 attttatggt gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga
     4201 cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac
     4261 agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg
     4321 aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata
     4381 ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     4441 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     4501 atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt
     4561 attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa
     4621 gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac
     4681 agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt
     4741 aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt
     4801 cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat
     4861 cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac
     4921 actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg
     4981 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
     5041 ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa
     5101 ctattaactg gcgaactact tactctagct tcccggcaac aattaataga ctggatggag
     5161 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
     5221 gataaatctg gagccggtga gcgtggaagc cgcggtatca ttgcagcact ggggccagat
     5281 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     5341 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac
     5401 caagtttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     5461 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     5521 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     5581 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     5641 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     5701 aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     5761 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     5821 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     5881 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     5941 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     6001 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     6061 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     6121 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     6181 ctggcctttt gctggccttt tgctcacatg t
//
