LOCUS       sequence-377336-        8139 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Cre-dependent expression of firefly luciferase, driven by a minimal 
            mouse Cry1 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8139)
  AUTHORS   Brancaccio M, Edwards MD, Patton AP, Smyllie NJ, Chesham JE, Maywood
            ES, Hastings MH
  TITLE     Cell-autonomous clock of astrocytes drives circadian behavior in 
            mammals.
  JOURNAL   Science. 2019 Jan 11;363(6423):187-192. doi: 
            10.1126/science.aat4104.
  PUBMED    30630934
REFERENCE   2  (bases 1 to 8139)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: "10.1126/science"; 
            journalName: "Science"; date: "2019-01-11- 11"; volume: "363"; 
            issue: "6423"; pages: "187-192"
COMMENT     Sequence Label: AAV.pCry1(408).DIO.Luciferase
FEATURES             Location/Qualifiers
     source          1..8139
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   417..546
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     protein_bind    complement(988..1021)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(1072..1105)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1114..2766)
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     2682..2700
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     protein_bind    2775..2808
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    2859..2892
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    2917..3505
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2970..2990)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3388..3399)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3537..4013
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3736..3755)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4053..4193
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4053..4182
                     /label=AAV2 ITR
     rep_origin      4268..4723
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4355..4374)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4565..4586
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4740..4759)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4859..4881
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4919..4937)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5005..5109
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5110..5178,5179..5970)
                     /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(5328..5347)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6141..6729
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     gap             7257..7393
                     /estimated_length=137
ORIGIN
        1 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
       61 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
      121 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
      181 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
      241 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
      301 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
      361 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgtcctg
      421 caggcagctg cgcgctcgct cgctcactga ggccgcccgg gcgtcgggcg acctttggtc
      481 gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccaactcc atcactaggg
      541 gttcctgcgg ccgcacgcgt cagaactatg cctcctcccc cttctcgcgg cctgtggccc
      601 ccgcgagcgg cctggggctc aggagacccg cagaggctgc gagtccggat cccacgcgag
      661 aactcaggtc gtgagcgccc agcgatcgca accggtcccg aagcaccggg cgcggcggcg
      721 cacgcggggg tctgagccag tgtagtaaac acacttcaga aacgtgaggt gccggtggtc
      781 acgtggggtg tatgccgccc aatgagaagc cgggggcggg gcggaggccg ctgacgcggc
      841 ggagtcaccc gggcagcctc gggaccggtg accggcaggc gagcgtcccg cggcctggag
      901 ctctagcccg cgctctccga gctcgctgcc cactccggga cgcacaaagc ttcgaattct
      961 gcagtcgact ctagagtcga ctccggaata acttcgtata ggatacttta tacgaagtta
     1021 tgcagaatgg tagctggatt gtagctgcta ttagcaatat gaaacctctt aataacttcg
     1081 tatagcatac attatacgaa gttatggcgc gccttacacg gcgatctttc cgcccttctt
     1141 ggcctttatg aggatctctc tgatttttct tgcgtcgagt tttccggtaa gacctttcgg
     1201 tacttcgtcc acaaacacaa ctcctccgcg caactttttc gcggttgtta cttgactggc
     1261 gacgtaatcc acgatctctt tttccgtcat cgtctttccg tgctccaaaa caacaacggc
     1321 ggcgggaagt tcaccggcgt catcgtcggg aagacctgcg acacctgcgt cgaagatgtt
     1381 ggggtgttgg agcaagatgg attccaattc agcgggagcc acctgatagc ctttgtactt
     1441 aatcagagac ttcaggcggt caacgatgaa gaagtgttcg tcttcgtccc agtaagctat
     1501 gtctccagaa tgtagccatc catccttgtc aatcaaggcg ttggtcgctt ccggattgtt
     1561 tacataaccg gacataatca taggacctct cacacacagt tcgcctcttt gattaacgcc
     1621 cagcgttttc ccggtatcca gatccacaac cttcgcttca aaaaatggaa caactttacc
     1681 gaccgcgccc ggtttatcat ccccctcggg tgtaatcaga atagctgatg tagtctcagt
     1741 gagcccatat ccttgcctga tacctggcag atggaacctc ttggcaaccg cttccccgac
     1801 ttccttagag aggggagcgc caccagaagc aatttcgtgt aaattagata aatcgtattt
     1861 gtcaatcaga gtgcttttgg cgaagaagga gaatagggtt ggcaccagca gcgcactttg
     1921 aatcttgtaa tcctgaaggc tcctcagaaa cagctcttct tcaaatctat acattaagac
     1981 gactcgaaat ccacatatca aatatccgag tgtagtaaac attccaaaac cgtgatggaa
     2041 tggaacaaca cttaaaatcg cagtatccgg aatgatttga ttgccaaaaa taggatctct
     2101 ggcatgcgag aatctcacgc aggcagttct atgaggcaga gcgacacctt taggcagacc
     2161 agtagatcca gaggagttca tgatcagtgc aattgtcttg tccctatcga aggactctgg
     2221 cacaaaatcg tattcattaa aaccgggagg tagatgagat gtgacgaacg tgtacatcga
     2281 ctgaaatccc tggtaatccg ttttagaatc catgataata attttttgga tgattgggag
     2341 ctttttttgc acgttcaaaa ttttttgcaa cccctttttg gaaacgaaca ccacggtagg
     2401 ctgcgaaatg cccatactgt tgagcaattc acgttcatta taaatgtcgt tcgcgggcgc
     2461 aactgcaact ccgataaata acgcgcccaa caccggcata aagaattgaa gagagttttc
     2521 actgcatacg acgattctgt gatttgtatt cagcccatat cgtttcatag cttctgccaa
     2581 ccgaacggac atttcgaagt actcagcgta agtgatgtcc acctcgatat gtgcatctgt
     2641 aaaagcaatt gttccaggaa ccagggcgta tctcttcata gccttatgca gttgctctcc
     2701 agcggttcca tcttccagcg gatagaatgg cgccgggcct ttctttatgt ttttggcgtc
     2761 ttccatgagc tagcataact tcgtataaag tatcctatac gaagttattt gccttaaccc
     2821 agaaattatc actgttattc tttagaatgg tgcaaagaat aacttcgtat aatgtatgct
     2881 atacgaagtt atgaattcga tatcaagctt atcgataatc aacctctgga ttacaaaatt
     2941 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct
     3001 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg
     3061 tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc
     3121 gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt
     3181 cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc
     3241 gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg
     3301 ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct atgttgccac ctggattctg
     3361 cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc
     3421 ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg
     3481 atctcccttt gggccgcctc cccgcatcga taccgagcgc tgctcgagag atctacgggt
     3541 ggcatccctg tgacccctcc ccagtgcctc tcctggccct ggaagttgcc actccagtgc
     3601 ccaccagcct tgtcctaata aaattaagtt gcatcatttt gtctgactag gtgtccttct
     3661 ataatattat ggggtggagg ggggtggtat ggagcaaggg gcaagttggg aagacaacct
     3721 gtagggcctg cggggtctat tgggaaccaa gctggagtgc agtggcacaa tcttggctca
     3781 ctgcaatctc cgcctcctgg gttcaagcga ttctcctgcc tcagcctccc gagttgttgg
     3841 gattccaggc atgcatgacc aggctcagct aatttttgtt tttttggtag agacggggtt
     3901 tcaccatatt ggccaggctg gtctccaact cctaatctca ggtgatctac ccaccttggc
     3961 ctcccaaatt gctgggatta caggcgtgaa ccactgctcc cttccctgtc cttctgattt
     4021 tgtaggtaac cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg agttggccac
     4081 tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc
     4141 gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct
     4201 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa
     4261 ccatagtacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     4321 gtgaccgcta cacttgccag cgccttagcg cccgctcctt tcgctttctt cccttccttt
     4381 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     4441 cgatttagtg ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt
     4501 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     4561 aatagtggac tcttgttcca aactggaaca acactcaact ctatctcggg ctattctttt
     4621 gatttataag ggattttgcc gatttcggtc tattggttaa aaaatgagct gatttaacaa
     4681 aaatttaacg cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt
     4741 acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac
     4801 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
     4861 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc
     4921 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca
     4981 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     5041 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     5101 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     5161 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     5221 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     5281 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     5341 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     5401 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     5461 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     5521 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     5581 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     5641 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     5701 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     5761 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     5821 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     5881 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     5941 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     6001 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     6061 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     6121 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     6181 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     6241 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     6301 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     6361 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     6421 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     6481 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     6541 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     6601 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     6661 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     6721 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggcmkwtyn ctrgmyktkk
     6781 ssymacrwkk cckgcrrsma gcwgsngcgc wcsswckyyy wcygcatcaa gtgttttggc
     6841 tctcaggccg aggcccacgg caagtatttg ggcaaggggt cgctggtatt cgtgcagggc
     6901 aagattcgga ataccaagta cgagaaggac ggccagacgg tctacgggac cgacttcatt
     6961 gccgataagg tggattatct ggacaccaag gcaccaggcg ggtcaaatca ggaataaggg
     7021 cacattgccc cggcgtgagt cggggcaatc ccgcaaggag ggtgaatgaa tcggacgttt
     7081 gaccggaagg catacaggca agaactgatc gacgcggggt tttccgccga ggatgccgaa
     7141 accatcgcaa gccgcaccgt catgcgtgcg ccccgcgaaa ccttccagtc cgtcggctcg
     7201 atggtccagc aagctacggc caagatcgag cgcgacagcg tgcaactggc tcccccnnnn
     7261 nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn
     7321 nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn
     7381 nnnnnnnnnn nnntggcaaa acaggtcagc gaggccaagc aggccgcgtt gctgaaacac
     7441 acgaagcagc agatcaagga aatgcagctt tccttgttcg atattgcgcc gtggccggac
     7501 acgatgcgag cgatgccaaa cgacacggcc cgctctgccc tgttcaccac gcgcaacaag
     7561 aaaatcccgc gcgaggcgct gcaaaacaag gtcattttcc acgtcaacaa ggacgtgaag
     7621 atcacctaca ccggcgtcga gctgcgggcc gacgatgacg aactggtgtg gcagcaggtg
     7681 ttggagtacg cgaagcgcac ccctatcggc gagccgatca ccttcacgtt ctacgagctt
     7741 tgccaggacc tgggctggtc gatcaatggc cggtattaca cgaaggccga ggaatgcctg
     7801 tcgcgcctac aggcgacggc gatgggcttc acgtccgacc gcgttgggca cctggaatcg
     7861 gtgtcgctgc tgcaccgctt ccgcgtcctg gaccgtggca agaaaacgtc ccgttgccag
     7921 gtcctgatcg acgaggaaat cgtcgtgctg tttgctggcg accactacac gaaattcata
     7981 tgggagaagt accgcaagct gtcgccgacg gcccgacgga tgttcgacta tttcagctcg
     8041 caccgggagc cgtacccgct caagctggaa accttccgcc tcatgtgcgg atcggattcc
     8101 acccgcgtga agaagtggcg cgagcaggtc ggcgaagcc
//
