LOCUS       sequence-334942-        5457 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CheRiff-T2A-mCherry construct expression in mammlian cells. pAAV 
            vector. Reporter plasmid..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5457)
  AUTHORS   Kaberniuk AA, Baloban M, Monakhov MV, Shcherbakova DM, Verkhusha VV
  TITLE     Single-component near-infrared optogenetic systems for gene 
            transcription regulation.
  JOURNAL   Nat Commun. 2021 Jun 23;12(1):3859. doi: 10.1038/s41467-021-24212-7.
  PUBMED    34162879
REFERENCE   2  (bases 1 to 5457)
  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: "Nat 
            Commun."; date: "2021-06-23"; volume: "12(1):3859. doi"; pages: " 
            10.1038/s41467-021-24212-7"
COMMENT     Sequence Label: pAAV-G12-CheRiff-T2A-mCherry
FEATURES             Location/Qualifiers
     source          1..5457
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        185..203
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     185..202
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     regulatory      511..520
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             517..1464
                     /codon_start=1
                     /product="blue light-activated optogenetic actuator derived
                     from Scherffelia dubia channelrhodopsin (Hochbaum et al., 
                     2014)"
                     /label=CheRiff
                     /translation="MGGAPAPDAHSAPPGNDSAGGSEYHAPAGYQVNPPYHPVHGYEEQ
                     CSSIYIYYGALWEQETARGFQWFAVFLSALFLAFYGWHAYKASVGWEEVYVCSVELIKV
                     ILEIYFEFTSPAMLFLYGGNITPWLRYAEWLLTCPVILIHLSNITGLSEAYNKRTMALL
                     VSDLGTICMGVTAALATGWVKWLFYCIGLVYGTQTFYNAGIIYVESYYIMPAGGCKKLV
                     LAMTAVYYSSWLMFPGLFIFGPEGMHTLSVAGSTIGHTIADLLSKNIWGLLGHFLRIKI
                     HEHIIMYGDIRRPVSSQFLGRKVDVLAFVTEEDKV"
     CDS             1552..1605
                     /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             1612..2319
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(1754..1772)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2022..2041
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(2389..2409)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     primer_bind     2591..2610
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   2720..2860
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2720..2849
                     /label=AAV2 ITR
     rep_origin      2935..3390
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3022..3041)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3232..3253
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3407..3426)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3526..3548
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3586..3604)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3672..3776
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3777..3845,3846..4637)
                     /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(3995..4014)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4808..5396
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5297..5316
                     /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 gcggccgaag atgctgcaca cttgtgggct agccttatgt atcatacaca
      181 tacgatttag gtgacactat agaactcgag cagctgaagc ttcggagtac tgtcctccga
      241 gcggagtact gtcctccgag cggagtactg tcctccgagt cgactctagc ggagtactgt
      301 cctccgagcg gagtactgtc ctccgagcgg agtactgtcc tccgagtcga ctctagcgga
      361 gtactgtcct ccgagcggag tactgtcctc cgagcggagt actgtcctcc gagtcgactc
      421 tagcggagta ctgtcctccg agcggagtac tgtcctccgg ctagtagcgg agactctaga
      481 gggtatataa tggatccccg ggtaccggtc gccaccatgg gcggagctcc tgctccagac
      541 gctcacagcg ccccacctgg aaacgattct gccggaggca gtgagtacca tgccccagct
      601 ggatatcaag tgaatccacc ctaccacccc gtgcatgggt atgaggaaca gtgcagctcc
      661 atctacatct actatggggc cctgtgggag caggaaacag ctaggggctt ccagtggttt
      721 gccgtgttcc tgtctgccct gtttctggct ttctacggct ggcacgccta taaggccagc
      781 gtgggatggg aggaagtgta cgtgtgctcc gtggagctga tcaaagtgat tctggagatc
      841 tatttcgagt tcaccagtcc tgctatgctg ttcctgtacg gagggaacat taccccatgg
      901 ctgagatatg ccgaatggct gctgacatgt cccgtgatcc tgattcatct gtctaacatc
      961 accggcctga gtgaggcata caataagcgg acaatggctc tgctggtgtc cgacctggga
     1021 actatttgca tgggagtgac agccgctctg gccactgggt gggtgaagtg gctgttttac
     1081 tgtatcggcc tggtgtatgg aacccagaca ttctacaacg ctggaatcat ctacgtggag
     1141 tcttactata tcatgcctgc cggcggctgt aagaaactgg tgctggccat gactgccgtg
     1201 tactattcta gttggctgat gtttcccggc ctgttcatct ttgggcctga aggcatgcac
     1261 accctgagcg tggctgggtc cactattggc cataccatcg ccgacctgct gtccaagaat
     1321 atttggggac tgctggggca cttcctgcgg atcaaaattc acgagcatat cattatgtac
     1381 ggcgatatca ggagaccagt gagctcccag tttctgggac gcaaggtgga cgtgctggcc
     1441 ttcgtgacag aggaagataa agtggcggcc gccaagagca ggatcaccag cgagggcgag
     1501 tacatccccc tggaccagat cgacatcaac gtggctcgag ataggcgcgc cgagggcaga
     1561 ggaagtctgc taacatgcgg tgacgtcgag gagaatcctg gcccaactag tgtgagcaag
     1621 ggcgaggagg ataacatggc catcatcaag gagttcatgc gcttcaaggt gcacatggag
     1681 ggctccgtga acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag
     1741 ggcacccaga ccgccaagct gaaggtgacc aagggtggcc ccctgccctt cgcctgggac
     1801 atcctgtccc ctcagttcat gtacggctcc aaggcctacg tgaagcaccc cgccgacatc
     1861 cccgactact tgaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc
     1921 gaggacggcg gcgtggtgac cgtgacccag gactcctccc tgcaggacgg cgagttcatc
     1981 tacaaggtga agctgcgcgg caccaacttc ccctccgacg gccccgtaat gcagaagaag
     2041 accatgggct gggaggcctc ctccgagcgg atgtaccccg aggacggcgc cctgaagggc
     2101 gagatcaagc agaggctgaa gctgaaggac ggcggccact acgacgctga ggtcaagacc
     2161 acctacaagg ccaagaagcc cgtgcagctg cccggcgcct acaacgtcaa catcaagttg
     2221 gacatcacct cccacaacga ggactacacc atcgtggaac agtacgaacg cgccgagggc
     2281 cgccactcca ccggcggcat ggacgagctg tacaagtaag aattccgctc gagataatca
     2341 acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt
     2401 tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc
     2461 tttcattttc tcctccttgt ataaatcctg gttagttctt gccacggcgg aactcatcgc
     2521 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt
     2581 gtttatttgt gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa
     2641 acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg agatgtggga
     2701 ggttttttaa agcggccgca ggaaccccta gtgatggagt tggccactcc ctctctgcgc
     2761 gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg
     2821 gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg ggcgcctgat gcggtatttt
     2881 ctccttacgc atctgtgcgg tatttcacac cgcatacgtc aaagcaacca tagtacgcgc
     2941 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     3001 ttgccagcgc cttagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     3061 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     3121 tacggcacct cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt gggccatcgc
     3181 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     3241 tgttccaaac tggaacaaca ctcaactcta tctcgggcta ttcttttgat ttataaggga
     3301 ttttgccgat ttcggtctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     3361 attttaacaa aatattaacg tttacaattt tatggtgcac tctcagtaca atctgctctg
     3421 atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg ccctgacggg
     3481 cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt
     3541 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc
     3601 tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt ggcacttttc
     3661 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     3721 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     3781 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     3841 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     3901 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     3961 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     4021 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     4081 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     4141 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     4201 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     4261 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     4321 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     4381 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     4441 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     4501 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     4561 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     4621 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     4681 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     4741 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     4801 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     4861 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     4921 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     4981 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     5041 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     5101 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     5161 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     5221 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     5281 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     5341 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     5401 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgt
//
