LOCUS       sequence-434417-        8100 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre-dependent expression of mCherry-tagged Chrimson; Optogenetics.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8100)
  AUTHORS   Soden ME, Chung AS, Cuevas B, Resnick JM, Awatramani R, Zweifel LS
  TITLE     Anatomic resolution of neurotransmitter-specific projections to the 
            VTA reveals diversity of GABAergic inputs.
  JOURNAL   Nat Neurosci. 2020 Aug;23(8):968-980. doi: 
            10.1038/s41593-020-0657-z. Epub 2020 Jun 15.
  PUBMED    32541962
REFERENCE   2  (bases 1 to 8100)
  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/s41593-020-0657-z"; journalName: "Nat Neurosci"; date: 
            "2020-08"; volume: "23"; issue: "8"; pages: "968-980"
COMMENT     Sequence Label: pAAV-CAG-FLEX-Chrimson-TdTomato
FEATURES             Location/Qualifiers
     source          1..8100
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     primer_bind     962..984
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1046..1065
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    complement(1116..1149)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(1200..1233)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(2697..3746)
                     /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      3743..3752
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3778..3811
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    3862..3895
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3920..4508
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3973..3993)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4391..4402)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4535..4552)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4541..4765
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        complement(4975..5304)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4989..5124
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5289..5309
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(5550..6138)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5630..5649)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6309..7100,7101..7169))
                     /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     6932..6951
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7170..7274)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7342..7360
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7398..7420)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7520..7539
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7753..7772
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        8062..8080
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     8062..8079
                     /label=SP6
                     /note="SP6 promoter, forward primer"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct
      181 aggtaccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      241 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      301 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      361 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      421 tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc atctcccccc
      481 cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca gcgatggggg
      541 cggggggggg gggggggggg cggggcgggg sgrggsggrg rggkgsggsg gcrgccaakc
      601 agagcggcgc gctccgaaag tttcctttta tggcgaggcg gcggcggcgg cggccctata
      661 aaaagcgaag cgcgcggcgg gcgggagtcg ctgcgcgctg ccttcgcccc gtgccccgct
      721 ccgccgccgc ctcgcgccgc ccgccccggc tctgactgac cgcgttactc ccacaggtga
      781 gcgggcggga cggcccttct cctccgggct gtaattagcg cttggtttaa tgacggcttg
      841 tttcttttct gtggctgcgt gaaagccttg aggggctccg ggagggccct ttgtgcgggg
      901 ggagcggctc ggggctgtcc gcggggggac ggctgccttc gggggggacg gggcagggcg
      961 gggttcggct tctggcgtgt gaccggcggc tctagagcct ctgctaacca tgttcatgcc
     1021 ttcttctttt tcctacagct cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt
     1081 ggcaaagaat tggatcctct agagtcgact ccggaataac ttcgtatagg atactttata
     1141 cgaagttatg cagaatggta gctggattgt agctgctatt agcaatatga aacctcttaa
     1201 taacttcgta tagcatacat tatacgaagt tatggcgcgc cgctagctca ttacttatac
     1261 agctcatcca tgccgtacag aaacagatgg tggcgtcctt cagacctctc atactgttcc
     1321 acaatggtgt agtcttcatt gtgggatgtg atatccagct tggtgtccac atagtaatag
     1381 ccgggcagct gcactggttt cttggccatg tagatggttt tgaactccac caggtagtgg
     1441 ccgccgtctt tcagcttcag ggcctggtgg atttcgccct tcagcactcc gtctctagga
     1501 tacagccgtt cagtgctggc ctcccacccc attgttttct tctgcatcac gggcccatcg
     1561 ggtggaaaat tggtgccgcg cattttgact ttgtagatca gagtgccatc ctgcagagag
     1621 gagtcctgtg tcacggtcac caggcctcca tcttcaaaat tcatcactct ctcccactta
     1681 aagccctctg ggaaagacag tttcttgtaa tcaggaatgt cagcgggatg cttgacatag
     1741 gccttacttc cgtacataaa ctgaggagac aggatatccc aagcgaaagg cagtggtcct
     1801 cccttggtca ctttcagctt ggcagtctgt gtgccctcgt aaggcctccc ttcgccctct
     1861 ccttcaatct caaattcgtg cccattcatg ctgccctcca tgcgcacctt aaacctcatg
     1921 aactctttga tcacagccat attgttatcc tcgcttgagg cggttccact agaccctgag
     1981 ccggtacttc cagtcccatg gcccaggaac agatggtgtc tcccctcgct ccgttcatac
     2041 tgctccacaa tagtgtagtc ctcgttgtgt gaagtgatat ccagttttgt gtccacgtaa
     2101 tagtagcctg gcagctgcac aggtttcttg gccatgtaga ttgtcttgaa ctccaccagg
     2161 taatgtcccc catctttcag cttcagagcc tggtggattt ctcctttcag caccccgtcg
     2221 cgtggataca gcctttcggt ggaggcctcc catcccatag ttttcttctg catcacaggt
     2281 ccatcagggg gaaaatttgt ccctctcatt ttcaccttgt agatcagggt cccatcctgc
     2341 agggagctgt cctgggtcac agtcaccagg cctccgtcct cgaagttcat cactctttcc
     2401 cacttaaacc cctcggggaa gctcagtttc ttataatcgg gaatgtcagc tggatgtttc
     2461 acataggcct tggagccgta cataaactgt gggctcagga tgtcccaagc gaatggcaga
     2521 ggcccgccct ttgtcacttt cagcttggct gtctgggtgc cctcgtaggg ccgtccctct
     2581 ccttctccct caatttcgaa ctcgtggccg ttcatgcttc cttccattct caccttaaac
     2641 cgcatgaact ctttgatcac ttcctcgccc ttactcactg ctactaccgg tgccgccact
     2701 gtgtcctcgt cctcctcctc cacgaactct tccacttcca cctcctcgcc tccgatctcc
     2761 atcttggtgg tcttccggat gtcgccgtgg atcaggatgt gctcgtggat cttgatcctc
     2821 aggtggtggg ccaggaaggt ccaaaactcc ttggcgatga tgtcgcagat gctgtggccg
     2881 atgctgttgg cgtaagggct cagcttcagc agtccttctg gtcccactgc ccagaggatt
     2941 gggtagctgc cccaagaggc gaagtaagcg taggccatca gcttcacgac catgcggcaa
     3001 tggcctttag gcacgctgtg gttggcttcc acgtagcact tggcggcctg gaagtacatg
     3061 tagccgccgt agatgcaaga cacgatatac agcagccact tgagccaatc ggtagccagt
     3121 cctgcggcca tgccgaacac gatcattccc acgcaagaca cgatcaggcc catggtccgc
     3181 ttgctgtagt cgttcttcag gccgctcagg ttgctcagtc tgatcaggat cacggggcaa
     3241 gacagcagcc actcgaagta gcgcaggcaa taggcgtggt tgccggtaga caggtacact
     3301 gtggcggggc tgctgaactc cttgaagatc tccagggtca cgaacagcac ctcgacgcaa
     3361 cagacgtaga cctcctccca accgcaagtg gccttccagg cgctgaagcc gtagaatgtc
     3421 agcagggcga tggcgatgct gaaagcaatc cactggcaga cctgggcgcc gatcttttct
     3481 cctggtgtgc ctctgctgtg cagccaagcc tgcttcttct ccactcccac gaccaggtag
     3541 ccgcctgtgg cctccacgaa gcaatatccg tatccggcgt cgctcagtcc gtaagaaggg
     3601 tcacaccacc actcggtgct gaagcactcg cctgtaggtg tcattcctcc acagcccatg
     3661 tcctcgtggt ggtaggggtt aggccaaggg ttgatgcctc cggcggcaaa cagagatctg
     3721 gtggcgctgc tgatcagctc agccatggtg gcgcggccgc tgcagaccgg tgctagcata
     3781 acttcgtata aagtatccta tacgaagtta tttgccttaa cccagaaatt atcactgtta
     3841 ttctttagaa tggtgcaaag aataacttcg tataatgtat gctatacgaa gttatgaatt
     3901 cgatatcaag cttatcgata atcaacctct ggattacaaa atttgtgaaa gattgactgg
     3961 tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta
     4021 tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct
     4081 gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt
     4141 tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac
     4201 tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg
     4261 ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaaatcatc
     4321 gtcctttcct tggctgctcg cctgtgttgc cacctggatt ctgcgcggga cgtccttctg
     4381 ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct
     4441 gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc
     4501 ctccccgcat cgataccgtc gactcgctga tcagcctcga ctgtgccttc tagttgccag
     4561 ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact
     4621 gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt
     4681 ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat
     4741 gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctcgact
     4801 agagcatggc tacgtagata agtagcatgg cgggttaatc attaactaca aggaacccct
     4861 agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc
     4921 aaaggtcgcc cgacgcccgg gcggcctcag tgagcgagcg agcgcgcaga gctttttgca
     4981 aaagcctagg cctccaaaaa agcctcctca ctacttctgg aatagctcag aggccgaggc
     5041 ggcctcggcc tctgcataaa taaaaaaaat tagtcagcca tggggcggag aatgggcgga
     5101 actgggcgga gttaggggcg ggatgggcgg agttaggggc gggactatgg ttgctgacta
     5161 attgagatgc atgctttgca tacttctgcc tgctggggag cctggggact ttccacacct
     5221 ggttgctgac taattgagat gcatgctttg catacttctg cctgctgggg agcctgggga
     5281 ctttccacac cctaactgac acacattcca cagctgcatt aatgaatcgg ccaacgcgcg
     5341 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     5401 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     5461 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     5521 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     5581 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     5641 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     5701 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     5761 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     5821 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     5881 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     5941 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     6001 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     6061 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     6121 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     6181 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     6241 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     6301 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     6361 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     6421 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     6481 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     6541 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     6601 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     6661 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     6721 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     6781 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     6841 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     6901 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     6961 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     7021 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     7081 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     7141 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     7201 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     7261 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga aaccattatt
     7321 atcatgacat taacctataa aaataggcgt atcacgaggc cctttcgtct cgcgcgtttc
     7381 ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg
     7441 taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt
     7501 cggggctggc ttaactatgc ggcatcagag cagattgtac tgagagtgca ccattcgacg
     7561 ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga ggccgttgag
     7621 caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg cccaacagtc ccccggccac
     7681 ggggcctgcc accataccca cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg
     7741 atcttcccca tcggtgatgt cggcgatata ggcgccagca accgcacctg tggcgccggt
     7801 gatgccggcc acgatgcgtc cggcgtagag gatctggcta gcgatgaccc tgctgattgg
     7861 ttcgctgacc atttccgggt gcgggacggc gttaccagaa actcagaagg ttcgtccaac
     7921 caaaccgact ctgacggcag tttacgagag agatgatagg gtctgcttca gtaagccaga
     7981 tgctacacaa ttaggcttgt acatattgtc gttagaacgc ggctacaatt aatacataac
     8041 cttatgtatc atacacatac gatttaggtg acactataga atacacggaa ttaattctag
//
