LOCUS       sequence-496081-        6889 bp    DNA     circular SYN 02-JAN-2026
DEFINITION  Reporter to measure ISR state-dependent protein synthesis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6889)
  AUTHORS   Helseth AR, Hernandez-Martinez R, Hall VL, Oliver ML, Turner BD, 
            Caffall ZF, Rittiner JE, Shipman MK, King CS, Gradinaru V, Gerfen C,
            Costa-Mattioli M, Calakos N
  TITLE     Cholinergic neurons constitutively engage the ISR for dopamine 
            modulation and skill learning in mice.
  JOURNAL   Science. 2021 Apr 23;372(6540). pii: 372/6540/eabe1931. doi: 
            10.1126/science.abe1931.
  PUBMED    33888613
REFERENCE   2  (bases 1 to 6889)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 2, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Science."; 
            date: "2021-04-23"; volume: "372(6540). pii: 372/6540/eabe1931. 
            doi"; pages: " 10.1126/science.abe1931"
COMMENT     Sequence Label: AAV-CAG-SPOTlight-U2GCR
FEATURES             Location/Qualifiers
     source          1..6889
                     /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"
     regulatory      1525..1534
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1630..1632,1633..1635,1636..1638,1639..2349)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MAVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLT
                     LKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKD
                     DGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGI
                     KVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVL
                     LEFVTAAGITLGMDELYK"
     CDS             join(1630..1632,1633..1635,1636..1638,1639..2349)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MAVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLT
                     LKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKD
                     DGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGI
                     KVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVL
                     LEFVTAAGITLGMDELYK"
     primer_bind     complement(1678..1699)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1939..1958)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             2368..2424
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2528..2557
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(3417..3435)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3879..3902
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     repeat_region   4247..4376
                     /label=AAV2 ITR
     rep_origin      complement(4613..5201)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4693..4712)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5372..6163,6164..6232))
                     /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     5995..6014
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6233..6337)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6363..6818)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6500..6521)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6712..6731
                     /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 ggcgcgcgcc aggcggggcg gggcggggcg aggggcgggg
      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 ccggctaggt
     1141 gtcccacccc gccttgtaag acaccggaaa ttcgtcaacg agcgatccct ccgcgctccg
     1201 gaaccagacc gcggctggtc gtcaacctat aaagtagtgc acttttctct catggggcct
     1261 ttaggacgat ctctaacgcc acagttaccc cccgagcaca gcggagaagg gttggggcgg
     1321 gcgagggcgt cagtgagagg cggtttggtg gcgctgcggt aggatcacgt gaccacagtg
     1381 ggaggatacg cggtgtgctg cgtccctggc cgaggctata aagggcgggt ttagggcgtg
     1441 ccgccgccat ttctgcttgc tgtctgccgg tttaagttgt gtgctcgggt gtccctttcc
     1501 tcttcccctc ccgcagggct tgcggccacc atggcgtatt agaggcagca gtgcctgcgg
     1561 cagcgttggc ctttgcagcg gcggcagcag caccaggctc tgcagcggca acccccaccg
     1621 gcctaagcca tggcggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
     1681 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
     1741 gacgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
     1801 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
     1861 gatcatatga agcagcacga cttcttcaag agtgcgatgc ccgaaggcta cgtccaggag
     1921 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
     1981 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     2041 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat aatggccgac
     2101 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     2161 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     2221 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     2281 gatcatatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     2341 ctgtacaagg gtagtggagg aagcggagct actaacttca gcctgctgaa gcaggctgga
     2401 gacgtggagg agaaccctgg acctgtggcg ctcttcacga aatccagcag cagtgttgct
     2461 gtaacggaca aagatacctt cgagttaagc acattcctcg aatccagcaa agccccacaa
     2521 catgactgaa caaaaactca tctcagaaga ggatctggtg gttagcaagg gcgaggaggt
     2581 gattaaggag tttatgcggt tcaaggtacg aatggaggga tctatgaacg gccacgaatt
     2641 tgaaatcgaa ggggagggag agggacgacc ttacgagggc actcagaccg ctaaattgaa
     2701 agtcacaaaa ggcggacctt tgccatttgc atgggacatc ctcagtcctc agtttatgta
     2761 cggcagtaag gcatatgtca aacatcccgc cgatattcca gattataaga aactttcctt
     2821 cccagaaggg ttcaaatggg agcgagtgat gaattttgaa gacggggggc tcgtaacagt
     2881 aacccaagac tctagccttc aagacggtac attgatttac aaagttaaga tgagaggtac
     2941 caacttcccc cccgacggtc ccgttatgca aaagaagaca atgggctggg aggccagcac
     3001 tgagcgactt tatcctaggg atggtgtcct caaaggcgag atccaccagg ctcttaaact
     3061 caaggacggc ggccattacc ttgttgaatt taaaactatc tatatggcta agaagcctgt
     3121 gcagttgcct ggttactatt acgtcgacac taagttggac ataacatccc ataatgaaga
     3181 ctacactatc gttgaacaat acgagcgatc agagggacgc catcacttgt tcctaggaca
     3241 cggtactggt agtactggta gtggcagctc cggcaccgcc tcctccgagg acaacaacat
     3301 ggccgtcatc aaagagttca tgcgcttcaa ggtgcgcatg gagggctcca tgaacggcca
     3361 cgagttcgag atcgagggcg agggcgaggg ccgcccctac gagggcaccc agaccgccaa
     3421 gctgaaggtg accaagggcg gccccctgcc cttcgcctgg gacatcctgt ccccccagtt
     3481 catgtacggc tccaaggcgt acgtgaagca ccccgccgac atccccgatt acaagaagct
     3541 gtccttcccc gagggcttca agtgggagcg cgtgatgaac ttcgaggacg gcggtctggt
     3601 gaccgtgacc caggactcct ccctgcagga cggcacgctg atctacaagg tgaagatgcg
     3661 cggcacaaac ttcccccccg acggccccgt aatgcagaag aagaccatgg gctgggaggc
     3721 ctccaccgag cgcctgtacc cccgcgacgg cgtgctgaag ggcgagatcc accaggccct
     3781 gaagctgaag gacggcggcc gctacctggt ggagttcaag accatctaca tggccaagaa
     3841 gcccgtgcaa ctgcccggct actactacgt ggacaccaag ctggacatca cctcccacaa
     3901 cgaggactac accatcgtgg aacagtacga gcgctccgag ggccgccacc acctgttcct
     3961 gtacggcatg gacgagctgt acaagtaata gggtagtcag gtgctttgtg cttgtacata
     4021 gtcttgtgtt gctgtgtttg ctgtaataaa ttattttgta gtgaaagtac ctatgtgggt
     4081 tttcagtgtc attcaattgc attcatttta tgtttcaggt tcagggggag atgtgggagg
     4141 ttttttaaag caagtaaaac ctctacaaat gtggtaaaat cgataaggat cttcctagag
     4201 catggctacg tagataagta gcatggcggg ttaatcatta actacaagga acccctagtg
     4261 atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag
     4321 gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc
     4381 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     4441 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     4501 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     4561 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     4621 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     4681 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     4741 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     4801 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     4861 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     4921 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     4981 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     5041 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     5101 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     5161 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     5221 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     5281 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     5341 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     5401 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     5461 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     5521 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     5581 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     5641 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     5701 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     5761 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     5821 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     5881 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     5941 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     6001 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     6061 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     6121 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     6181 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     6241 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     6301 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     6361 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     6421 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
     6481 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
     6541 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     6601 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     6661 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     6721 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     6781 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
     6841 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccag
//
