LOCUS       sequence-457821-        5354 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 5354)
  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 5354)
  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-BC7-pA
FEATURES             Location/Qualifiers
     source          1..5354
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     promoter        441..717
                     /label=chicken beta-actin promoter
     primer_bind     996..1018
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1080..1099
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1138..1833
                     /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(1271..1289)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1539..1558
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    1866..2454
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1919..1939)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2337..2348)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2578..2699
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2589..2608)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2643..2662
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(3078..3666)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3158..3177)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3837..4628,4629..4697))
                     /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     4460..4479
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4698..4802)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4828..5283)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4965..4986)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5177..5196
                     /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 gcgcgcgcca ggcggggcgg ggcggggcga ggggcggggc
      601 ggggcgaggc ggagaggtgc ggcggcagcc aatcagagcg gcgcgctccg aaagtttcct
      661 tttatggcga ggcggcggcg gcggcggccc tataaaaagc gaagcgcgcg gcgggcggga
      721 gtcgctgcgc gctgccttcg ccccgtgccc cgctccgccg ccgcctcgcg ccgcccgccc
      781 cggctctgac tgaccgcgtt actcccacag gtgagcgggc gggacggccc ttctcctccg
      841 ggctgtaatt agcgcttggt ttaatgacgg cttgtttctt ttctgtggct gcgtgaaagc
      901 cttgaggggc tccgggaggg ccctttgtgc ggggggagcg gctcggggct gtccgcgggg
      961 ggacggctgc cttcgggggg gacggggcag ggcggggttc ggcttctggc gtgtgaccgg
     1021 cggctctaga gcctctgcta accatgttca tgccttcttc tttttcctac agctcctggg
     1081 caacgtgctg gttattgtgc tgtctcatca ttttggcaaa gaattggatc cgaattcatg
     1141 gtgagcaagg gcgaggcagt gatcaaggag ttcatgcggt tcaaggtgca catggagggc
     1201 tccatgaacg gccacgagtt cgagatcgag ggcgagggcg agggccgccc ctacgagggc
     1261 acccagaccg ccaagctgaa ggtgaccaag ggtggccccc tgcccttctc ctgggacatc
     1321 ctgtcccctc agttcatgta cggctccagg gccttcacca agcaccccgc cgacatcccc
     1381 gactactata agcagtcctt ccccgagggc ttcaagtggg accgcgtgat gaacttcgag
     1441 gacggcggcg ccgtgaccgt gacccaggac acctccctgg aggacggcac cctgatctac
     1501 aaggtgaagc tccgcggcac caacttccct cctgacggcc ccgtaatgca gaagaagaca
     1561 atgggctggg aagcgtccac cgagcggttg taccccgagg acggcgtgct gaagggcgac
     1621 attaagatgg ccctgcgcct gaaggacggc gggcgctacc tggcggactt caagaccacc
     1681 tacaaggcca agaagcccgt gcagatgccc ggcgcctaca acgtggaccg caagttggac
     1741 atcacctccc acaacgagga ctacaccgtg gtggaacagt acgaacgctc cgagggccgc
     1801 cactccaccg gcggcatgga cgagctgtac aagtaaaagc tttctagagg cgcgagctta
     1861 tcgataatca acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg
     1921 ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt
     1981 cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg
     2041 agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc
     2101 ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc
     2161 tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc
     2221 ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc
     2281 tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg
     2341 ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cgtcttccgc
     2401 gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcgcggcc
     2461 gcacttggag gtgttgcagg tggtgcacaa gttcctccac aagatgtact tggagaacca
     2521 gctcgacttg gtcttggaca acgtcgtgtt ctagcaggtc cattcgagca gacatgataa
     2581 gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt
     2641 gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta
     2701 acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggagatgtgg gaggtttttt
     2761 aaagcaagta aaacctctac aaatgtggta aaatcgataa ggatcttcct agagcatggc
     2821 tacgtagata agtagcatgg cgggttaatc attaactaca aggaacccct agtgatggag
     2881 ttggccactc cctctctgcg cgctcgctcg ctcactgact cgctgcgctc ggtcgttcgg
     2941 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     3001 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     3061 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     3121 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     3181 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     3241 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     3301 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     3361 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     3421 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     3481 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct
     3541 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     3601 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     3661 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     3721 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
     3781 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac
     3841 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
     3901 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
     3961 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
     4021 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
     4081 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
     4141 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
     4201 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
     4261 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
     4321 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
     4381 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
     4441 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
     4501 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
     4561 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
     4621 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
     4681 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
     4741 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     4801 cgcacatttc cccgaaaagt gccacctaaa ttgtaagcgt taatattttg ttaaaattcg
     4861 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc
     4921 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga
     4981 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
     5041 atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag
     5101 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
     5161 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
     5221 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
     5281 cgtcccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt
     5341 cgctattacg ccag
//
