LOCUS       sequence-229628-        6546 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector for sparse and bright labeling in a cell-type specific 
            manner.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6546)
  AUTHORS   Lin R, Wang R, Yuan J, Feng Q, Zhou Y, Zeng S, Ren M, Jiang S, Ni H,
            Zhou C, Gong H, Luo M
  TITLE     Cell-type-specific and projection-specific brain-wide reconstruction
            of single neurons.
  JOURNAL   Nat Methods. 2018 Nov 19. pii: 10.1038/s41592-018-0184-y. doi: 
            10.1038/s41592-018-0184-y.
  PUBMED    30455464
REFERENCE   2  (bases 1 to 6546)
  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 
            Methods. 2018 Nov 19. pii: 10.1038/s41592-018-0184-y. doi: 
            10.1038/s41592-018-0184-y."
COMMENT     Sequence Label: pAAV-TRE-vDIO-mScarlet-IRES-tTA
FEATURES             Location/Qualifiers
     source          1..6546
                     /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        148..462
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    156..174
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    192..210
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    227..245
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    263..281
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    299..317
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    334..352
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    370..388
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     435..459
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             complement(join(559..567,568..603,604..606,607..642,
                     643..645,646..681,682..687,688..1305))
                     /codon_start=1
                     /product="improved tetracycline-controlled transactivator"
                     /label=tTA-Advanced
                     /note="In the Tet-Off(R) system, tTA-Advanced binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the absence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGGPADALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     misc_feature    1314..1866
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    1316..1889
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1476..1495)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     1705..1722
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     CDS             complement(1894..2589)
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(2169..2188)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2438..2456
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     regulatory      2586..2595
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    2686..3274
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2739..2759)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3157..3168)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3293..3769
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3492..3511)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3809..3949
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3809..3938
                     /label=AAV2 ITR
     rep_origin      4024..4479
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4111..4130)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4321..4342
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4496..4515)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4615..4637
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4675..4693)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4761..4865
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4866..4934,4935..5726)
                     /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(5084..5103)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5897..6485
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6386..6405
                     /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 gcgtctcgag tttactccct atcagtgata gagaacgtat
      181 gtcgagttta ctccctatca gtgatagaga acgatgtcga gtttactccc tatcagtgat
      241 agagaacgta tgtcgagttt actccctatc agtgatagag aacgtatgtc gagtttactc
      301 cctatcagtg atagagaacg tatgtcgagt ttatccctat cagtgataga gaacgtatgt
      361 cgagtttact ccctatcagt gatagagaac gtatgtcgag gtaggcgtgt acggtgggag
      421 gcctatataa gcagagctcg tttagtgaac cgtcagatcg cctggagaat tcttaattaa
      481 tcaatttctg agaagtgtct ttctcggaaa ttgaggtcaa tttctgagaa ctgtcattct
      541 cggaaattga ggcgcgcctt acccggggag catgtcaagg tcaaaatcgt caagagcgtc
      601 agcaggcagc atatcaaggt caaagtcgtc aagggcatcg gctgggagca tgtctaagtc
      661 aaaatcgtca agggcgtcgg ccggcccgcc gctttcgcac tttagctgtt tctccaggcc
      721 acatatgatt agttccaggc cgaaaaggaa ggcaggttcg gctccctgat ggtcgaacag
      781 ctcaattgct tgtctcagaa gtgggggcat agaatcggtg gtaggtgtct ctctttcctc
      841 ttttgctact tgatgctcct gatcctccaa tacgcagccc agtgtaaagt ggcccacggc
      901 ggacagagcg tacagtgcgt tctccaggga gaagccttgc tgacacagga acgcgagctg
      961 attttccagg gtttcgtact gtttctctgt tgggcgggtg ccgagatgca ctttagcccc
     1021 gtcgcgatgt gagaggagag cacagcggaa tgacttggcg ttgttccgca gaaagtcttg
     1081 ccatgactcg ccttccaggg ggcagaagtg ggtatgatgc ctgtccagca tctcgattgc
     1141 cagggcatcg agcagggccc gcttgttctt cacgtgccag tacagggtag gctgctcaac
     1201 tcccagcttt tgagcgagtt tccttgtcgt caggccttcg ataccgactt cattgagtaa
     1261 ttccagagca gagtttatga ctttgctctt gtccagtcta gacattgtgg ccatattatc
     1321 atcgtgtttt tcaaaggaaa accacgtccc cgtggttcgg ggggcctaga cgttttttta
     1381 acctcgacta aacacatgta aagcatgtgt accgaggccc cagatcagat cccatacaat
     1441 ggggtacctt ctgggcatcc ttcagcccct tgttgaatac gcttgaggag agccatttga
     1501 ctctttccac aactatccaa ctcacaacgt ggcactgggg ttgtgccgcc tttgcaggtg
     1561 tatcttatac acgtggcttt tggccgcaga ggcacctgtc gccaggtggg gggttccgct
     1621 gcctgcaaag ggtcgctaca gacgttgttt gtcttcaaga agcttccaga ggaactgctt
     1681 ccttcacgac attcaacaga ccttgcattc ctttggcgag aggggaaaga cccctaggaa
     1741 tgctcgtcaa gaagacaggg ccaggtttcc gggccctcac attgccaaaa gacggcaata
     1801 tggtggaaaa taacatatag acaaacgcac accggcctta ttccaagcgg cttcggccag
     1861 taacgttagg ggggggggag ggagaggggc ttacttgtac agctcatcca tgccgccggt
     1921 ggagtggcgg ccctcggagc gttcgtactg ttccaccacg gtgtagtcct cgttgtggga
     1981 ggtgatgtcc aacttgcggt cgacgttgta ggcgccgggc atctgcacgg gcttcttggc
     2041 cttgtaggtg gtcttgaagt ccgccaggta gcggccgccg tccttcaggc gcagggccat
     2101 cttaatgtcg cccttcagca cgccgtcctc ggggtacaac cgctcggtgg acgcttccca
     2161 gcccattgtc ttcttctgca ttacggggcc gtcaggaggg aagttggtgc cgcggagctt
     2221 caccttgtag atcagggtgc cgtcctccag ggaggtgtcc tgggtcacgg tcacggcgcc
     2281 gccgtcctcg aagttcatca cgcgctccca cttgaagccc tcggggaagg actgcttata
     2341 gtagtcgggg atgtcggcgg ggtgcttggt gaaggccctg gagccgtaca tgaactgagg
     2401 ggacaggatg tcccaggaga agggcagggg gccacccttg gtcaccttca gcttggcggt
     2461 ctgggtgccc tcgtaggggc ggccctcgcc ctcgccctcg atctcgaact cgtggccgtt
     2521 catggagccc tccatgtgca ccttgaaccg catgaactcc ttgatcactg cctcgccctt
     2581 gctcaccatg gtggctagct caatttccga gaaagacact tctcagaaat tgagctaatt
     2641 caatttccga gaatgacagt tctcagaaat tgaaagctta tcgataatca acctctggat
     2701 tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt
     2761 ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc
     2821 tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg
     2881 caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc
     2941 accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa
     3001 ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat
     3061 tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgccta tgttgccacc
     3121 tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt
     3181 ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag
     3241 acgagtcgga tctccctttg ggccgcctcc ccgcatcgat accgagatct acgggtggca
     3301 tccctgtgac ccctccccag tgcctctcct ggccctggaa gttgccactc cagtgcccac
     3361 cagccttgtc ctaataaaat taagttgcat cattttgtct gactaggtgt ccttctataa
     3421 tattatgggg tggagggggg tggtatggag caaggggcaa gttgggaaga caacctgtag
     3481 ggcctgcggg gtctattggg aaccaagctg gagtgcagtg gcacaatctt ggctcactgc
     3541 aatctccgcc tcctgggttc aagcgattct cctgcctcag cctcccgagt tgttgggatt
     3601 ccaggcatgc atgaccaggc tcagctaatt tttgtttttt tggtagagac ggggtttcac
     3661 catattggcc aggctggtct ccaactccta atctcaggtg atctacccac cttggcctcc
     3721 caaattgctg ggattacagg cgtgaaccac tgctcccttc cctgtccttc tgattttgta
     3781 ggtaaccacg tgcggaccga gcggccgcag gaacccctag tgatggagtt ggccactccc
     3841 tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc
     3901 tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg
     3961 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat
     4021 agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     4081 ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct tcctttctcg
     4141 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     4201 ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg
     4261 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     4321 gtggactctt gttccaaact ggaacaacac tcaactctat ctcgggctat tcttttgatt
     4381 tataagggat tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat
     4441 ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa
     4501 tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc
     4561 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
     4621 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg
     4681 tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg
     4741 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     4801 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     4861 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     4921 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     4981 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     5041 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     5101 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     5161 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     5221 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     5281 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     5341 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     5401 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     5461 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     5521 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     5581 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     5641 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     5701 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     5761 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     5821 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     5881 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     5941 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     6001 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     6061 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     6121 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     6181 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     6241 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     6301 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     6361 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     6421 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     6481 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     6541 acatgt
//
