LOCUS       sequence-439275-        7168 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 7168)
  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 7168)
  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-BC6-pA
FEATURES             Location/Qualifiers
     source          1..7168
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     primer_bind     999..1021
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1083..1102
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             join(2800..2802,2803..3516)
                     /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             2800..3513
                     /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(2830..2858)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     3453..3473
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     misc_feature    3534..4122
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3587..3607)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4005..4016)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4265..4386
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4276..4295)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4330..4349
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(4892..5480)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4972..4991)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5651..6442,6443..6511))
                     /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     6274..6293
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6512..6616)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6642..7097)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6779..6800)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6991..7010
                     /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 gcggggcgcg gggggggggg gggggcgggg cgaggggcgg
      601 ggcggggcga ggcggagagg tgcggcggca gccaatcaga gcggcgcgct ccgaaagttt
      661 ccttttatgg cgaggcggcg gcggcggcgg ccctataaaa agcgaagcgc gcggcgggcg
      721 ggagtcgctg cgcgctgcct tcgccccgtg ccccgctccg ccgccgcctc gcgccgcccg
      781 ccccggctct gactgaccgc gttactccca caggtgagcg ggcgggacgg cccttctcct
      841 ccgggctgta attagcgctt ggtttaatga cggcttgttt cttttctgtg gctgcgtgaa
      901 agccttgagg ggctccggga gggccctttg tgcgggggga gcggctcggg gctgtccgcg
      961 gggggacggc tgccttcggg ggggacgggg cagggcgggg ttcggcttct ggcgtgtgac
     1021 cggcggctct agagcctctg ctaaccatgt tcatgccttc ttctttttcc tacagctcct
     1081 gggcaacgtg ctggttattg tgctgtctca tcattttggc aaagaattgg atccgaattc
     1141 actagcatga cctccagggt gtactccaga gacaggactc tcaaaccacg gaaggcagcc
     1201 tcgggaacct tctggtggct agggagcggc tggtaccaat ttgttacttt gatttcttgg
     1261 ctgaatgtct ttcttcttac caggtgcctt cgaaatattt gcaaggtttt tgtcttgctc
     1321 ctcccactcc tacttttact aggtgctggt gtctccctgt ggggccaggg aaacttcttc
     1381 tcactcctac cagtgctgaa ctggacggcc atgcagccaa cacagagggt ggacgattcc
     1441 aagggcatgc atagacctgg ccctcttccc ccgagcccac ctccaaaggt tgatcacaag
     1501 gcttcccagt ggcctcagga gagtgacatg gggcagaagg tagcttcttt gagtgcgcag
     1561 tgccacaacc atgatgagag acttgcagag ctgacagtcc tgcttcagaa actacagata
     1621 cgggtagacc aagtggatga cggcagggaa gggctgtcac tgtgggtcaa gaatgtggtt
     1681 ggacagcacc tgcaggagat gggcaccata gaaccacctg atgctaagac tgacttcatg
     1741 actttccacc atgaccatga agtgcgtctc tccaacttgg aagatgttct tagaaaactg
     1801 acagaaaaat ctgaggctat ccagaaggag ctggaagaaa ccaagctgaa agcaggcagc
     1861 agggatgaag agcagcccct ccttgaccgt gtgcagcacc tagaactgga actgaacctg
     1921 ttgaagtcac agctgtcaga ctggcagcat ctgaagacca gctgtgagca ggctggggcc
     1981 cgcatccagg agactgtgca gctcatgttc tctgaggatc agcagggcgg ttccctcgag
     2041 tggctattag agaagctttc ttctcggttc gtgagcaagg atgagctgca ggtgctctta
     2101 catgaccttg agctgaaact gctgcagaat atcacacacc acatcaccgt gacaggacag
     2161 gccccgacat ccgaggctat tgtgtctgcc gtgaatcagg cagggatttc aggaatcaca
     2221 gaagcgcaag cacatatcat tgtgaacaat gctctgaagc tgtactccca agacaagacg
     2281 gggatggtgg actttgctct ggagtctgga ggtggcagca tcctaagcac tcggtgctct
     2341 gagacctatg agaccaagac ggcactgctg agcctgtttg gggtcccact gtggtacttc
     2401 tcacagtcac ctcgagtggt gatccagccc gacatctacc cagggaattg ctgggcgttc
     2461 aaaggttccc aggggtacct ggtggtgcgg ttgtccatga agatctaccc aaccacattc
     2521 accatggaac acattccaaa gacactatca cccactggta acatctccag tgcccccaaa
     2581 gactttgcag tctatggact ggaaacggag tatcaagaag aggggcagcc tctgggacgg
     2641 ttcacctatg accaggaagg agactcactc cagatgttcc acacactgga aagacctgac
     2701 caagccttcc agatagtaga gctccgggtc ctgtccaact ggggccaccc tgagtacact
     2761 tgcctctacc ggttccgagt ccacggagag cccatccaga tgagcaaagg agaagaactt
     2821 ttcactggag ttgtcccaat tcttgttgaa ttagatggtg atgttaatgg gcacaaattt
     2881 tctgtccgtg gagagggtga aggtgatgct acaaacggaa aactcaccct taaatttatt
     2941 tgcactactg gaaaactacc tgttccatgg ccaacacttg tcactactct gacctatggt
     3001 gttcaatgct tttcccgtta tccggatcac atgaaacggc atgacttttt caagagtgcc
     3061 atgcccgaag gttatgtaca ggaacgcact atatctttca aagatgacgg gacctacaag
     3121 acgcgtgctg aagtcaagtt tgaaggtgat acccttgtta atcgtatcga gttaaaaggt
     3181 attgatttta aagaagatgg aaacattctc ggacacaaac tcgagtacaa ctttaactca
     3241 cacaatgtat acatcacggc agacaaacaa aagaatggaa tcaaagctaa cttcaaaatt
     3301 cgccacaacg ttgaagatgg ttccgttcaa ctagcagacc attatcaaca aaatactcca
     3361 attggcgatg gccctgtcct tttaccagac aaccattacc tgtcgacaca atctgtcctt
     3421 tcgaaagatc ccaacgaaaa gcgtgaccac atggtccttc ttgagtttgt aactgctgct
     3481 gggattacac atggcatgga tgagctctac aaataaggcg cgagcttatc gataatcaac
     3541 ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta
     3601 cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt
     3661 tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg
     3721 ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg
     3781 gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca
     3841 cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca
     3901 ctgacaattc cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctgtg
     3961 ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag
     4021 cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc
     4081 gccctcagac gagtcggatc tccctttggg ccgcctcccc gcatcgatac cgtcgacccg
     4141 ggcggccgca cgtcaacgtg gacgtcctcg tgcaggtcgt gtaggtggtg ttggtcaacc
     4201 agcagaacta ccaggagctg gagctccagt tgcacttcaa ggagtagcat tcgagcagac
     4261 atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc
     4321 tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa
     4381 caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga gatgtgggag
     4441 gttttttaaa gcaagtaaaa cctctacaaa tgtggtaaaa tcgataagga tcttcctaga
     4501 gcatggctac gtagataagt agcatggcgg gttaatcatt aactacaagg aacccctagt
     4561 gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg cccgggcttt
     4621 gcccgggcgg cctcagtgag cgagcgagcg cgcagctgca ttaatgaatc ggccaacgcg
     4681 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
     4741 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     4801 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     4861 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     4921 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     4981 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     5041 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     5101 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     5161 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     5221 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     5281 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     5341 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     5401 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     5461 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     5521 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     5581 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     5641 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
     5701 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
     5761 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
     5821 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
     5881 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
     5941 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
     6001 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
     6061 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
     6121 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
     6181 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
     6241 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
     6301 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
     6361 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
     6421 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
     6481 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
     6541 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
     6601 aaataggggt tccgcgcaca tttccccgaa aagtgccacc taaattgtaa gcgttaatat
     6661 tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga
     6721 aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc
     6781 agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac
     6841 cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc
     6901 gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg
     6961 gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag
     7021 ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc
     7081 gccgctacag ggcgcgtccc attcgccatt caggctgcgc aactgttggg aagggcgatc
     7141 ggtgcgggcc tcttcgctat tacgccag
//
