LOCUS       sequence-151865-        5927 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses GFP from the CBA promoter. Cloned into an Adeno-Associated
            Virus backbone for packaging into AAV..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5927)
  AUTHORS   Borchardt EK, Vandoros LA, Huang M, Lackey PE, Marzluff WF, Asokan A
  TITLE     Controlling mRNA stability and translation with the CRISPR 
            endoribonuclease Csy4.
  JOURNAL   RNA. 2015 Nov;21(11):1921-30. doi: 10.1261/rna.051227.115. Epub 2015
            Sep 9.
  PUBMED    26354771
REFERENCE   2  (bases 1 to 5927)
  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"; doi: 
            "10.1261/rna.051227.115"; journalName: "RNA"; date: "2015-11"; 
            volume: "21"; issue: "11"; pages: "1921-30"
COMMENT     Sequence Label: TR-GFP
FEATURES             Location/Qualifiers
     source          1..5927
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      120..129
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(126..128,129..131,132..845)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(126..128,129..131,132..842)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(171..192)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(432..451)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     779..800
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    874..995
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(885..904)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     939..958
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      1346..1801
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1433..1452)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1643..1664
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1868..1887)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1987..2009
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2047..2065)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2133..2237
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2238..2306,2307..3098)
                     /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     complement(2456..2475)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3269..3857
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3758..3777
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        4292..4671
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4673..4950
                     /label=chicken beta-actin promoter
     primer_bind     5893..5915
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
ORIGIN
        1 cgggctgtaa ttagctgagc aagaggtaag ggtttaaggg atggttggtt ggtggggtat
       61 taatgtttaa ttacctggag cacctgcctg aaatcacttt ttttcaggtt ggaccggtcg
      121 ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
      181 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
      241 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
      301 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
      361 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
      421 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
      481 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
      541 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
      601 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
      661 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
      721 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
      781 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
      841 agtaactcga gcggccgctt cgagcagaca tgataagata cattgatgag tttggacaaa
      901 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
      961 tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta
     1021 tgtttcaggt tcagggggag atgtgggagg ttttttaaag caagtaaaac ctctacaaat
     1081 gtggtaaaat cgataaggat ctaggaaccc ctagtgatgg agttggccac tccctctctg
     1141 cgcgctcgct cgctcactga ggccgcccgg gcaaagcccg ggcgtcgggc gacctttggt
     1201 cgcccggcct cagtgagcga gcgagcgcgc agagagggag tggccaaccc cccccccccc
     1261 ccccctgcag cctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg
     1321 cgtagcctga atggcgaatg gcgcgacgcg ccctgtagcg gcgcattaag cgcggcgggt
     1381 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     1441 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     1501 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     1561 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     1621 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     1681 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     1741 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gtttacaatt
     1801 tcctgatgcg ctattttctc cttacgcatc tgtgcggtat ttcacaccgc atatggtgca
     1861 ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac
     1921 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     1981 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac
     2041 gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt
     2101 agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     2161 aaatactttc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     2221 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     2281 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     2341 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     2401 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
     2461 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact
     2521 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     2581 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     2641 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     2701 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     2761 agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg
     2821 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
     2881 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgcggat aaatctggag
     2941 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc
     3001 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
     3061 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
     3121 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     3181 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     3241 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     3301 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     3361 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc
     3421 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     3481 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     3541 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     3601 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     3661 attgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     3721 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     3781 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     3841 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     3901 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
     3961 ccgcctttga gtgagctgat accgctgggg gggggggggg gggttggcca ctccctctct
     4021 gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc
     4081 ccgggcggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact ccatcactag
     4141 gggttcctag atcttcaata ttggccatta gccatattat tcattggtta tatagcataa
     4201 atcaatattg gatattggcc attgcatacg ttgtatctat atcataatat gtacatttat
     4261 attggctcat gtccaatatg accgccatgt tggcattgat tattgactag ttattaatag
     4321 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
     4381 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
     4441 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
     4501 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tccgccccct
     4561 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttacgg
     4621 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtcgaggtg
     4681 agccccacgt tctgcttcac tctccccatc tcccccccct ccccaccccc aattttgtat
     4741 ttatttattt tttaattatt ttgtgcagcg atgggggcgg gggggggggg ggggcgcgcg
     4801 ccaggcgggg cggggcgggg cgaggggcgg ggcggggcga ggcggagagg tgcggcggca
     4861 gccaatcaga gcggcgcgct ccgaaagttt ccttttatgg cgaggcggcg gcggcggcgg
     4921 ccctataaaa agcgaagcgc gcggcgggcg ggagtcgctg cgacgctgcc ttcgccccgt
     4981 gccccgctcc gccgccgcct cgcgccgccc gccccggctc tgactgaccg cgttactccc
     5041 acaggtgagc gggcgggacg gcccttctcc tccgggctgt aattagcgct tggtttaatg
     5101 acggcttgtt tcttttctgt ggctgcgtga aagccttgag gggctccggg agggcccttt
     5161 gtgcgggggg gagcggctcg gggggtgcgt gcgtgtgtgt gtgcgtgggg agcgccgcgt
     5221 gcggcccgcg ctgcccggcg gctgtgagcg ctgcgggcgc ggcgcggggc tttgtgcgct
     5281 ccgcagtgtg cgcgagggga gcgcggccgg gggcggtgcc ccgcggtgcg gggggggctg
     5341 cgaggggaac aaaggctgcg tgcggggtgt gtgcgtgggg gggtgagcag ggggtatggg
     5401 cgcggcggtc gggctgtaac ccccccctgc acccccctcc ccgagttgct gagcacggcc
     5461 cggcttcggg tgcggggctc cgtacggggc gtggcgcggg gctcgccgtg ccgggcgggg
     5521 ggtggcggca ggtgggggtg ccgggcgggg cggggccgcc tcgggccggg gagggctcgg
     5581 gggaggggcg cggcggcccc cggagcgccg gcggctgtcg aggcgcggcg agccgcagcc
     5641 attgcctttt atggtaatcg tgcgagaggg cgcagggact tactttgtcc caaatctgtg
     5701 cggagccgaa atctgggagg cgccgccgca ccccctctag cgggcgcggg gcgaagcggt
     5761 gcggcgccgg caggaaggaa atgggcgggg agggccttcg tgcgtcgccg cgccgccgtc
     5821 cccttctccc tctccagcct cggggctgtc cgcgggggga cggctgcctt cgggggggac
     5881 ggggcagggc ggggttcggc ttctggcgtg tgaccggcgg ctctaga
//
