LOCUS       sequence-211671-        7339 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of full length mouse CRY1 mutant K31R.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7339)
  AUTHORS   Yoo SH, Mohawk JA, Siepka SM, Shan Y, Huh SK, Hong HK, Kornblum I, 
            Kumar V, Koike N, Xu M, Nussbaum J, Liu X, Chen Z, Chen ZJ, Green 
            CB, Takahashi JS
  TITLE     Competing E3 ubiquitin ligases govern circadian periodicity by 
            degradation of CRY in nucleus and cytoplasm.
  JOURNAL   Cell. 2013 Feb 28;152(5):1091-105. doi: 10.1016/j.cell.2013.01.055.
  PUBMED    23452855
REFERENCE   2  (bases 1 to 7339)
  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: 
            "10.1016/j.cell.2013.01"; journalName: "Cell"; date: "2013-02-28- 
            28"; volume: "152"; issue: "5"; pages: "1091-105"
COMMENT     Sequence Label: mcry1K31R HA tag
FEATURES             Location/Qualifiers
     source          1..7339
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        109..488
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        489..692
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     643..663
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     737..756
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        737..755
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2672..2698
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2717..2746
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2762..2779
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2802..2819)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2808..3032
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3078..3506
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3165..3184)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3375..3396
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3515..3535)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3520..3849
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3700..3835
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3762..3781
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3916..4710
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3970..3989)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4580..4599
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4886..5007
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4923..4942)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4977..4996
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5056..5072)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5056..5072)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5069..5091)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5080..5096
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5104..5110,5111..5128,5129..5134))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5149..5170
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5287..5304)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5458..6046)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5538..5557)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6217..7008,7009..7077))
                     /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     6840..6859
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7078..7182)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7257..7276)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat ctgcttaggg
       61 ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat
      121 tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt
      181 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
      241 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
      301 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
      361 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
      421 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
      481 ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac
      541 ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc
      601 aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc
      661 gtgtacggtg ggaggtctat ataagcagag ctctctggct aactagagaa cccactgctt
      721 actggcttat cgaaattaat acgactcact atagggagac ccaagctggc tagttaagct
      781 tggtaccgag ctcggatcca ctagtccagt gtggtggaat tctgcagata tccagcacag
      841 tggcggccgc accatgggtg ttaatgccgt ccattggttt agaagaggcc tccgtctgca
      901 tgacaatccg gctctgcgcg aatgtatcca gggtgctgac acgatcaggt gcgtttacat
      961 tcttgaccct tggtttgcag gcagcagcaa cgtaggcatc aatagatggc gtttccttct
     1021 ccagtgtctg gaggatttgg atgccaatct gcgtaggttg aactcccgtt tgtttgttat
     1081 tcgcggccag cctgctgacg tgtttccccg cctgttccgc gaatggaaca ttacacgtct
     1141 gagcattgag tacgactctg agccatttgg tcgtgaacgc gatgccgcta tccgcagact
     1201 cgccacagag gctggagtcg aggttatcgt gagaataagc cacacactgt acgacctcga
     1261 caggatcata gagctcaacg gtggtcaacc tccgctgact tacaggcgct ttcagaccct
     1321 cgtaagcaga atggagccgc tcgaaatgcc tgccgataca atcacctccg acgtgatcgg
     1381 ccgctgcatg acgccacttt cggacgacca cgatgagaga tacggagtac ctagcttgga
     1441 agagctcgga tttgacactg atggactgtc aagtgcggtg tggccaggtg gtgaaaccga
     1501 ggcattgacc aggcttgagc gtcaccttga acgccgtgct tgggttgcca acttcgaaag
     1561 gccacgtatg aatgcgaact ctctcctggc ttctcctact ggactctcgc cctaccttag
     1621 gtttggctgc ctgagttgtc gtttgttcta tttccgcctg accgacctct atagacgcgt
     1681 gcgtcgtaac agttccccac cactctcatt gtatggtcag ctcttgtgga gggaattctt
     1741 ctacactgcg gcgacaaaca atcccagatt cgatcgtatg gaaggaaatc ccatctgcgt
     1801 tcagatccct tgggacagaa accccgaggc gttggcacgt tgggctgaag gacgcaccgg
     1861 tttcccctgg atagacgcca tcatgaccca gctgcgccaa gagggttgga ttcaccatct
     1921 ggctaggcac gcagtcgcct gtttcctcac cagaggcgac ctttggattt catgggagga
     1981 gggcatgcgt gtattcgaag agttgctcct tgatgctgat tggtccatta acgcaggctc
     2041 ttggatgtgg ttgtcttgct cctcgttctt ccaacaattc ttccactgct attgcccagt
     2101 gggtttcgga aggaggaccg acccgaacgg tgactacatc cgccgctatc tgccggtcct
     2161 taggggcttc cctgcgcgtt acatctacga cccctggaat gctcccgaag gaattcagcg
     2221 tgtcgctcgc tgcctgattg gtgtcaacta ccctcgtcct atggtcaatc atgccgaggc
     2281 ctcccgcttg aacatagaga gaatgcgtca aatctaccag caactctctc gttatagagg
     2341 attgggactg ctggcatcgg tgccaagtaa ctcgaacggt aacggcggac tgatgggata
     2401 tgctcccggc gagaacgtgc cctcgtgttc ctcaagtggt aatggaggcc ttatgggtta
     2461 cgctcccggt gaaaacgtgc catcttgctc tggtggaaac tgctcccaag gctccggcat
     2521 tttgcactac gcccacggtg actctcagca gactcattct ctgaggcagg gtcgttcatc
     2581 agctggcacg ggcttgtcca gcggcaggag gccctcccaa gaggaagatg cacagtcagt
     2641 cggtccgaga gtacaaagac aaagcagtaa ctatccctat gacgtcccgg actatgcata
     2701 atctagaggg cccttcgaac aaaaactcat ctcagaagag gatctgaata tgcataccgg
     2761 tcatcatcac catcaccatt gagtttaaac ccgctgatca gcctcgactg tgccttctag
     2821 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     2881 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca
     2941 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag
     3001 caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg
     3061 ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     3121 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
     3181 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     3241 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
     3301 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     3361 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     3421 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
     3481 gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga
     3541 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     3601 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     3661 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     3721 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     3781 gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     3841 ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca agagacagga
     3901 tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     3961 gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc
     4021 gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt
     4081 gccctgaatg aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt
     4141 ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc
     4201 gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc
     4261 atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac
     4321 caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag
     4381 gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag
     4441 gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat
     4501 atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg
     4561 gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa
     4621 tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc
     4681 ttctatcgcc ttcttgacga gttcttctga gcgggactct ggggttcgcg aaatgaccga
     4741 ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag
     4801 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     4861 catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata
     4921 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     4981 tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag
     5041 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     5101 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     5161 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     5221 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     5281 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     5341 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     5401 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     5461 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     5521 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     5581 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     5641 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     5701 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     5761 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     5821 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     5881 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     5941 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     6001 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     6061 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     6121 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     6181 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     6241 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     6301 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     6361 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     6421 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     6481 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     6541 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     6601 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     6661 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     6721 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     6781 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     6841 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     6901 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     6961 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     7021 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     7081 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     7141 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     7201 gccacctgac gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca
     7261 atctgctctg atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc
     7321 gctgagtagt gcgcgagca
//
