LOCUS       sequence-439001-        7166 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 7166)
  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 7166)
  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-BC11-pA
FEATURES             Location/Qualifiers
     source          1..7166
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     promoter        441..718
                     /label=chicken beta-actin promoter
     primer_bind     997..1019
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1081..1100
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             join(2798..2800,2801..3514)
                     /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             2798..3511
                     /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(2828..2856)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     3451..3471
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     misc_feature    3532..4120
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3585..3605)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4003..4014)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4263..4384
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4274..4293)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4328..4347
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(4890..5478)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4970..4989)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5649..6440,6441..6509))
                     /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     6272..6291
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6510..6614)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6640..7095)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6777..6798)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6989..7008
                     /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 ggggcgcgcc gggggggggg gggggggggg gggggggggg
      601 cggggcgagg cggagaggtg cggcggcagc caatcagagc ggcgcgctcc gaaagtttcc
      661 ttttatggcg aggcggcggc ggcggcggcc ctataaaaag cgaagcgcgc ggcgggcggg
      721 agtcgctgcg cgctgccttc gccccgtgcc ccgctccgcc gccgcctcgc gccgcccgcc
      781 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      841 gggctgtaat tagcgcttgg tttaatgacg gcttgtttct tttctgtggc tgcgtgaaag
      901 ccttgagggg ctccgggagg gccctttgtg cggggggagc ggctcggggc tgtccgcggg
      961 gggacggctg ccttcggggg ggacggggca gggcggggtt cggcttctgg cgtgtgaccg
     1021 gcggctctag agcctctgct aaccatgttc atgccttctt ctttttccta cagctcctgg
     1081 gcaacgtgct ggttattgtg ctgtctcatc attttggcaa agaattggat ccgaattcac
     1141 tagcatgacc tccagggtgt actccagaga caggactctc aaaccacgga aggcagcctc
     1201 gggaaccttc tggtggctag ggagcggctg gtaccaattt gttactttga tttcttggct
     1261 gaatgtcttt cttcttacca ggtgccttcg aaatatttgc aaggtttttg tcttgctcct
     1321 cccactccta cttttactag gtgctggtgt ctccctgtgg ggccagggaa acttcttctc
     1381 actcctacca gtgctgaact ggacggccat gcagccaaca cagagggtgg acgattccaa
     1441 gggcatgcat agacctggcc ctcttccccc gagcccacct ccaaaggttg atcacaaggc
     1501 ttcccagtgg cctcaggaga gtgacatggg gcagaaggta gcttctttga gtgcgcagtg
     1561 ccacaaccat gatgagagac ttgcagagct gacagtcctg cttcagaaac tacagatacg
     1621 ggtagaccaa gtggatgacg gcagggaagg gctgtcactg tgggtcaaga atgtggttgg
     1681 acagcacctg caggagatgg gcaccataga accacctgat gctaagactg acttcatgac
     1741 tttccaccat gaccatgaag tgcgtctctc caacttggaa gatgttctta gaaaactgac
     1801 agaaaaatct gaggctatcc agaaggagct ggaagaaacc aagctgaaag caggcagcag
     1861 ggatgaagag cagcccctcc ttgaccgtgt gcagcaccta gaactggaac tgaacctgtt
     1921 gaagtcacag ctgtcagact ggcagcatct gaagaccagc tgtgagcagg ctggggcccg
     1981 catccaggag actgtgcagc tcatgttctc tgaggatcag cagggcggtt ccctcgagtg
     2041 gctattagag aagctttctt ctcggttcgt gagcaaggat gagctgcagg tgctcttaca
     2101 tgaccttgag ctgaaactgc tgcagaatat cacacaccac atcaccgtga caggacaggc
     2161 cccgacatcc gaggctattg tgtctgccgt gaatcaggca gggatttcag gaatcacaga
     2221 agcgcaagca catatcattg tgaacaatgc tctgaagctg tactcccaag acaagacggg
     2281 gatggtggac tttgctctgg agtctggagg tggcagcatc ctaagcactc ggtgctctga
     2341 gacctatgag accaagacgg cactgctgag cctgtttggg gtcccactgt ggtacttctc
     2401 acagtcacct cgagtggtga tccagcccga catctaccca gggaattgct gggcgttcaa
     2461 aggttcccag gggtacctgg tggtgcggtt gtccatgaag atctacccaa ccacattcac
     2521 catggaacac attccaaaga cactatcacc cactggtaac atctccagtg cccccaaaga
     2581 ctttgcagtc tatggactgg aaacggagta tcaagaagag gggcagcctc tgggacggtt
     2641 cacctatgac caggaaggag actcactcca gatgttccac acactggaaa gacctgacca
     2701 agccttccag atagtagagc tccgggtcct gtccaactgg ggccaccctg agtacacttg
     2761 cctctaccgg ttccgagtcc acggagagcc catccagatg agcaaaggag aagaactttt
     2821 cactggagtt gtcccaattc ttgttgaatt agatggtgat gttaatgggc acaaattttc
     2881 tgtccgtgga gagggtgaag gtgatgctac aaacggaaaa ctcaccctta aatttatttg
     2941 cactactgga aaactacctg ttccatggcc aacacttgtc actactctga cctatggtgt
     3001 tcaatgcttt tcccgttatc cggatcacat gaaacggcat gactttttca agagtgccat
     3061 gcccgaaggt tatgtacagg aacgcactat atctttcaaa gatgacggga cctacaagac
     3121 gcgtgctgaa gtcaagtttg aaggtgatac ccttgttaat cgtatcgagt taaaaggtat
     3181 tgattttaaa gaagatggaa acattctcgg acacaaactc gagtacaact ttaactcaca
     3241 caatgtatac atcacggcag acaaacaaaa gaatggaatc aaagctaact tcaaaattcg
     3301 ccacaacgtt gaagatggtt ccgttcaact agcagaccat tatcaacaaa atactccaat
     3361 tggcgatggc cctgtccttt taccagacaa ccattacctg tcgacacaat ctgtcctttc
     3421 gaaagatccc aacgaaaagc gtgaccacat ggtccttctt gagtttgtaa ctgctgctgg
     3481 gattacacat ggcatggatg agctctacaa ataaggcgcg agcttatcga taatcaacct
     3541 ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg
     3601 ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc
     3661 attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt
     3721 gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac tggttggggc
     3781 attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc tattgccacg
     3841 gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact
     3901 gacaattccg tggtgttgtc ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt
     3961 gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct caatccagcg
     4021 gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc
     4081 cctcagacga gtcggatctc cctttgggcc gcctccccgc atcgataccg tcgacccggg
     4141 cggccgcaga acttgcacca cgacatcgag ctccaggtcg tcgtcatcat cgtcatggag
     4201 gacttggtcg acgacgtcta gtaggtggac gagaagaagc agttggtttc gagcagacat
     4261 gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt
     4321 tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca
     4381 agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggaga tgtgggaggt
     4441 tttttaaagc aagtaaaacc tctacaaatg tggtaaaatc gataaggatc ttcctagagc
     4501 atggctacgt agataagtag catggcgggt taatcattaa ctacaaggaa cccctagtga
     4561 tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc
     4621 ccgggcggcc tcagtgagcg agcgagcgcg cagctgcatt aatgaatcgg ccaacgcgcg
     4681 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     4741 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     4801 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     4861 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     4921 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     4981 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     5041 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     5101 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     5161 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     5221 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     5281 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     5341 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     5401 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     5461 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     5521 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     5581 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     5641 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     5701 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     5761 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     5821 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     5881 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     5941 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     6001 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     6061 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     6121 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     6181 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     6241 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     6301 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     6361 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     6421 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     6481 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     6541 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     6601 ataggggttc cgcgcacatt tccccgaaaa gtgccaccta aattgtaagc gttaatattt
     6661 tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa
     6721 tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag
     6781 tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg
     6841 tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga
     6901 ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg
     6961 gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg
     7021 cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc
     7081 cgctacaggg cgcgtcccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg
     7141 tgcgggcctc ttcgctatta cgccag
//
