LOCUS       sequence-457818-        7186 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of Sun1GFP for nuclei 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 7186)
  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 7186)
  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-Sun1-GFP-WPRE-BC4-pA
FEATURES             Location/Qualifiers
     source          1..7186
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     promoter        441..716
                     /label=chicken beta-actin promoter
     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             join(2796..2798,2799..3512)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             2796..3509
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Nager et al., 2011)"
                     /label=superfolder GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(2826..2854)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     3449..3469
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     misc_feature    3530..4118
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3583..3603)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4001..4012)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4261..4382
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4272..4291)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4326..4345
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(4910..5498)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4990..5009)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5669..6460,6461..6529))
                     /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     6292..6311
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6530..6634)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6660..7115)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6797..6818)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7009..7028
                     /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 sgcgcgccag gcggggcggg gcggggcgag gggcggggcg
      601 gggcgaggcg 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 gaattcacta
     1141 gcatgacctc cagggtgtac tccagagaca ggactctcaa accacggaag gcagcctcgg
     1201 gaaccttctg gtggctaggg agcggctggt accaatttgt tactttgatt tcttggctga
     1261 atgtctttct tcttaccagg tgccttcgaa atatttgcaa ggtttttgtc ttgctcctcc
     1321 cactcctact tttactaggt gctggtgtct ccctgtgggg ccagggaaac ttcttctcac
     1381 tcctaccagt gctgaactgg acggccatgc agccaacaca gagggtggac gattccaagg
     1441 gcatgcatag acctggccct cttcccccga gcccacctcc aaaggttgat cacaaggctt
     1501 cccagtggcc tcaggagagt gacatggggc agaaggtagc ttctttgagt gcgcagtgcc
     1561 acaaccatga tgagagactt gcagagctga cagtcctgct tcagaaacta cagatacggg
     1621 tagaccaagt ggatgacggc agggaagggc tgtcactgtg ggtcaagaat gtggttggac
     1681 agcacctgca ggagatgggc accatagaac cacctgatgc taagactgac ttcatgactt
     1741 tccaccatga ccatgaagtg cgtctctcca acttggaaga tgttcttaga aaactgacag
     1801 aaaaatctga ggctatccag aaggagctgg aagaaaccaa gctgaaagca ggcagcaggg
     1861 atgaagagca gcccctcctt gaccgtgtgc agcacctaga actggaactg aacctgttga
     1921 agtcacagct gtcagactgg cagcatctga agaccagctg tgagcaggct ggggcccgca
     1981 tccaggagac tgtgcagctc atgttctctg aggatcagca gggcggttcc ctcgagtggc
     2041 tattagagaa gctttcttct cggttcgtga gcaaggatga gctgcaggtg ctcttacatg
     2101 accttgagct gaaactgctg cagaatatca cacaccacat caccgtgaca ggacaggccc
     2161 cgacatccga ggctattgtg tctgccgtga atcaggcagg gatttcagga atcacagaag
     2221 cgcaagcaca tatcattgtg aacaatgctc tgaagctgta ctcccaagac aagacgggga
     2281 tggtggactt tgctctggag tctggaggtg gcagcatcct aagcactcgg tgctctgaga
     2341 cctatgagac caagacggca ctgctgagcc tgtttggggt cccactgtgg tacttctcac
     2401 agtcacctcg agtggtgatc cagcccgaca tctacccagg gaattgctgg gcgttcaaag
     2461 gttcccaggg gtacctggtg gtgcggttgt ccatgaagat ctacccaacc acattcacca
     2521 tggaacacat tccaaagaca ctatcaccca ctggtaacat ctccagtgcc cccaaagact
     2581 ttgcagtcta tggactggaa acggagtatc aagaagaggg gcagcctctg ggacggttca
     2641 cctatgacca ggaaggagac tcactccaga tgttccacac actggaaaga cctgaccaag
     2701 ccttccagat agtagagctc cgggtcctgt ccaactgggg ccaccctgag tacacttgcc
     2761 tctaccggtt ccgagtccac ggagagccca tccagatgag caaaggagaa gaacttttca
     2821 ctggagttgt cccaattctt gttgaattag atggtgatgt taatgggcac aaattttctg
     2881 tccgtggaga gggtgaaggt gatgctacaa acggaaaact cacccttaaa tttatttgca
     2941 ctactggaaa actacctgtt ccatggccaa cacttgtcac tactctgacc tatggtgttc
     3001 aatgcttttc ccgttatccg gatcacatga aacggcatga ctttttcaag agtgccatgc
     3061 ccgaaggtta tgtacaggaa cgcactatat ctttcaaaga tgacgggacc tacaagacgc
     3121 gtgctgaagt caagtttgaa ggtgataccc ttgttaatcg tatcgagtta aaaggtattg
     3181 attttaaaga agatggaaac attctcggac acaaactcga gtacaacttt aactcacaca
     3241 atgtatacat cacggcagac aaacaaaaga atggaatcaa agctaacttc aaaattcgcc
     3301 acaacgttga agatggttcc gttcaactag cagaccatta tcaacaaaat actccaattg
     3361 gcgatggccc tgtcctttta ccagacaacc attacctgtc gacacaatct gtcctttcga
     3421 aagatcccaa cgaaaagcgt gaccacatgg tccttcttga gtttgtaact gctgctggga
     3481 ttacacatgg catggatgag ctctacaaat aaggcgcgag cttatcgata atcaacctct
     3541 ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct
     3601 atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat
     3661 tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt
     3721 caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat
     3781 tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc
     3841 ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga
     3901 caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg cctgtgttgc
     3961 cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga
     4021 ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc
     4081 tcagacgagt cggatctccc tttgggccgc ctccccgcat cgataccgtc gacccgggcg
     4141 gccgctggac cagtagttcg tcctgatctt gcagttcttg ttgttgaacc agctgttcta
     4201 gctcgtcatg cagatggacg agttctagga ctacttcgtc ttcatttcga gcagacatga
     4261 taagatacat tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta
     4321 tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag
     4381 ttaacaacaa caattgcatt cattttatgt ttcaggttca gggggagatg tgggaggttt
     4441 tttaaagcaa gtaaaacctc tacaaatgtg gtaaaatcga taaggatctt cctagagcat
     4501 ggctacgtag ataagtagca tggcgggtta atcattaact acaaggaacc cctagtgatg
     4561 gagttggcca ctccctctct gcgcgctcgc tcgctcactg aggccgcsck ggctsmscmc
     4621 kgmsgccgcc cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg cagctgcatt
     4681 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     4741 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     4801 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     4861 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     4921 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     4981 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     5041 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     5101 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     5161 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     5221 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     5281 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     5341 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     5401 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     5461 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     5521 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     5581 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     5641 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     5701 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     5761 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     5821 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     5881 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     5941 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     6001 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     6061 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     6121 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     6181 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     6241 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     6301 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     6361 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     6421 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     6481 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     6541 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     6601 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccaccta
     6661 aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt
     6721 ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat
     6781 agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa
     6841 cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta
     6901 atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc
     6961 ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc
     7021 gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac
     7081 acccgccgcg cttaatgcgc cgctacaggg cgcgtcccat tcgccattca ggctgcgcaa
     7141 ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccag
//
