LOCUS       sequence-436181-        5480 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of oScarlet for cell isolation and Projection-TAG BC for 
            multiplex projection tracing.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5480)
  AUTHORS   Yang L, Liu F, Hahm H, Okuda T, Li X, Zhang Y, Kalyanaraman V, 
            Heitmeier MR, Samineni VK
  TITLE     Projection-TAGs enable multiplex projection tracing and multi-modal 
            profiling of projection neurons.
  JOURNAL   Nat Commun. 2025 Jul 1;16(1):5557. doi: 10.1038/s41467-025-60360-w.
  PUBMED    40593543
REFERENCE   2  (bases 1 to 5480)
  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: "2025-07-1"; volume: "16(1):5557. doi"; pages: " 
            10.1038/s41467-025-60360-w"
COMMENT     Sequence Label: pAV-CAG-oScarlet-WPRE-BC2-pA
FEATURES             Location/Qualifiers
     source          1..5480
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     primer_bind     995..1017
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1079..1098
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1137..1832
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWDRVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(1270..1288)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1538..1557
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    1865..2453
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1918..1938)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2336..2347)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2577..2698
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2588..2607)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2642..2661
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(3204..3792)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3284..3303)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3963..4754,4755..4823))
                     /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     4586..4605
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4824..4928)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4954..5409)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5091..5112)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5303..5322
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct
      181 aggaagagta ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
      241 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
      301 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
      361 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
      421 agtcatcgct attaccatgg tcgaggtgag ccccacgttc tgcttcactc tccccatctc
      481 ccccccctcc ccacccccaa ttttgtattt atttattttt taattatttt gtgcagcgat
      541 gggggcgggg gggggggggg ggcgcgccrg gggggggggg gggggggggg ggggggggsg
      601 gggsgrggcg gagaggtgcg gcggcagcca atcagagcgg cgcgctccga aagtttcctt
      661 ttatggcgag gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag
      721 tcgctgcgcg ctgccttcgc cccgtgcccc gctccgccgc cgcctcgcgc cgcccgcccc
      781 ggctctgact gaccgcgtta ctcccacagg tgagcgggcg ggacggccct tctcctccgg
      841 gctgtaatta gcgcttggtt taatgacggc ttgtttcttt tctgtggctg cgtgaaagcc
      901 ttgaggggct ccgggagggc cctttgtgcg gggggagcgg ctcggggctg tccgcggggg
      961 gacggctgcc ttcggggggg acggggcagg gcggggttcg gcttctggcg tgtgaccggc
     1021 ggctctagag cctctgctaa ccatgttcat gccttcttct ttttcctaca gctcctgggc
     1081 aacgtgctgg ttattgtgct gtctcatcat tttggcaaag aattggatcc gaattcatgg
     1141 tgagcaaggg cgaggcagtg atcaaggagt tcatgcggtt caaggtgcac atggagggct
     1201 ccatgaacgg ccacgagttc gagatcgagg gcgagggcga gggccgcccc tacgagggca
     1261 cccagaccgc caagctgaag gtgaccaagg gtggccccct gcccttctcc tgggacatcc
     1321 tgtcccctca gttcatgtac ggctccaggg ccttcaccaa gcaccccgcc gacatccccg
     1381 actactataa gcagtccttc cccgagggct tcaagtggga ccgcgtgatg aacttcgagg
     1441 acggcggcgc cgtgaccgtg acccaggaca cctccctgga ggacggcacc ctgatctaca
     1501 aggtgaagct ccgcggcacc aacttccctc ctgacggccc cgtaatgcag aagaagacaa
     1561 tgggctggga agcgtccacc gagcggttgt accccgagga cggcgtgctg aagggcgaca
     1621 ttaagatggc cctgcgcctg aaggacggcg ggcgctacct ggcggacttc aagaccacct
     1681 acaaggccaa gaagcccgtg cagatgcccg gcgcctacaa cgtggaccgc aagttggaca
     1741 tcacctccca caacgaggac tacaccgtgg tggaacagta cgaacgctcc gagggccgcc
     1801 actccaccgg cggcatggac gagctgtaca agtaaaagct ttctagaggc gcgagcttat
     1861 cgataatcaa cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt
     1921 tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc
     1981 ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga
     2041 gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc
     2101 cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct
     2161 ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg
     2221 gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct
     2281 gctcgcctgt gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc
     2341 cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg
     2401 tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcgcggccg
     2461 ctgaaccact tctaggtgga cttggtctag atcatgtagt acttctagct gttggtgatc
     2521 gacctcaagt tcaacaactt gatcctcttg gtcatggtgg attcgagcag acatgataag
     2581 atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg
     2641 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa
     2701 caacaacaat tgcattcatt ttatgtttca ggttcagggg gagatgtggg aggtttttta
     2761 aagcaagtaa aacctctaca aatgtggtaa aatcgataag gatcttccta gagcatggct
     2821 acgtagataa gtagcatggc gggttaatca ttaactacaa ggaaccccta gtgatggagt
     2881 tggccactcc ctctctgcgc gctcgctcgc tcactgaggc cgcccgggct ttgcccgggc
     2941 ggcctcagtg agcgagcgag cgcgcagctg cattaatgaa tcggccaacg cgcggggaga
     3001 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     3061 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     3121 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     3181 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     3241 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     3301 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     3361 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     3421 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     3481 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     3541 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     3601 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     3661 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     3721 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     3781 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     3841 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     3901 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     3961 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     4021 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     4081 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     4141 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     4201 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     4261 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     4321 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     4381 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     4441 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     4501 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     4561 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     4621 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     4681 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     4741 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     4801 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     4861 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     4921 gttccgcgca catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa
     4981 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     5041 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     5101 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     5161 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
     5221 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     5281 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
     5341 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     5401 agggcgcgtc ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg
     5461 cctcttcgct attacgccag
//
