LOCUS       sequence-310822-        7000 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresssion and protein purification of full length RUVBL2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7000)
  AUTHORS   Ju D, Zhang W, Yan J, Zhao H, Li W, Wang J, Liao M, Xu Z, Wang Z, 
            Zhou G, Mei L, Hou N, Ying S, Cai T, Chen S, Xie X, Lai L, Tang C, 
            Park N, Takahashi JS, Huang N, Qi X, Zhang EE
  TITLE     Chemical perturbations reveal that RUVBL2 regulates the circadian 
            phase in mammals.
  JOURNAL   Sci Transl Med. 2020 May 6;12(542). pii: 12/542/eaba0769. doi: 
            10.1126/scitranslmed.aba0769.
  PUBMED    32376767
REFERENCE   2  (bases 1 to 7000)
  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: "Sci Transl 
            Med."; date: "2020-05-6"; volume: "12(542). pii: 12/542/eaba0769. 
            doi"; pages: " 10.1126/scitranslmed.aba0769"
COMMENT     Sequence Label: pET-28a RUVBL2 Full Length
FEATURES             Location/Qualifiers
     source          1..7000
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(1..1083)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     1045..1064
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1084..1161)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1367..1386
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1470..1489
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1470..1488
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1489..1513
                     /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)."
     RBS             1528..1550
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1569..1586
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1596..1613
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1626..1919
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
                     RRLMEAFAKRQGKEMDSLRFLYDGIRIQADQTPEDLDMEDNDIIEAHREQIGG"
     CDS             3334..3351
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3404..3422)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3418..3465
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3502..3957
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3589..3608)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3799..3820
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(4049..4864)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(4049..4864)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     4772..4791
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4856..4875
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4986..5574
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5475..5494
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5728..5745
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5760..5899
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5825..5844)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5985..6007
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6001..6192)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
       61 cgcgcgggga gaggcggttt gcgtattggg cgccagggtg gtttttcttt tcaccagtga
      121 gacgggcaac agctgattgc ccttcaccgc ctggccctga gagagttgca gcaagcggtc
      181 cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg gcgggatata
      241 acatgagctg tcttcggtat cgtcgtatcc cactaccgag atatccgcac caacgcgcag
      301 cccggactcg gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg caaccagcat
      361 cgcagtggga acgatgccct cattcagcat ttgcatggtt tgttgaaaac cggacatggc
      421 actccagtcg ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga gatatttatg
      481 ccagccagcc agacgcagac gcgccgagac agaacttaat gggcccgcta acagcgcgat
      541 ttgctggtga cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt cttcatggga
      601 gaaaataata ctgttgatgg gtgtctggtc agagacatca agaaataacg ccggaacatt
      661 agtgcaggca gcttccacag caatggcatc ctggtcatcc agcggatagt taatgatcag
      721 cccactgacg cgttgcgcga gaagattgtg caccgccgct ttacaggctt cgacgccgct
      781 tcgttctacc atcgacacca ccacgctggc acccagttga tcggcgcgag atttaatcgc
      841 cgcgacaatt tgcgacggcg cgtgcagggc cagactggag gtggcaacgc caatcagcaa
      901 cgactgtttg cccgccagtt gttgtgccac gcggttggga atgtaattca gctccgccat
      961 cgccgcttcc actttttccc gcgttttcgc agaaacgtgg ctggcctggt tcaccacgcg
     1021 ggaaacggtc tgataagaga caccggcata ctctgcgaca tcgtataacg ttactggttt
     1081 cacattcacc accctgaatt gactctcttc cgggcgctat catgccatac cgcgaaaggt
     1141 tttgcgccat tcgatggtgt ccgggatctc gacgctctcc cttatgcgac tcctgcatta
     1201 ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat
     1261 gcaaggagat ggcgcccaac agtcccccgg ccacggggcc tgccaccata cccacgccga
     1321 aacaagcgct catgagcccg aagtggcgag cccgatcttc cccatcggtg atgtcggcga
     1381 tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt
     1441 agaggatcga gatctcgatc ccgcgaaatt aatacgactc actatagggg aattgtgagc
     1501 ggataacaat tcccctctag aaataatttt gtttaacttt aagaaggaga tataccatgg
     1561 gcagcagcca tcatcatcat catcacagca gcggcctggt gccgcgcggc agccatatgg
     1621 ctagcatgtc ggactcagaa gtcaatcaag aagctaagcc agaggtcaag ccagaagtca
     1681 agcctgagac tcacatcaat ttaaaggtgt ccgatggatc ttcagagatc ttcttcaaga
     1741 tcaaaaagac cactccttta agaaggctga tggaagcgtt cgctaaaaga cagggtaagg
     1801 aaatggactc cttaagattc ttgtacgacg gtattaggat tcaagctgat cagacccctg
     1861 aagatttgga catggaggat aacgatatta ttgaggctca cagagaacag attggtggat
     1921 ccatggcaac cgttacagcc acaaccaaag tcccggagat ccgtgatgta acaaggattg
     1981 agcgaatcgg tgcccactcc cacatccggg gactggggct ggacgatgcc ttggagcctc
     2041 ggcaggcttc gcaaggcatg gtgggtcagc tggcggcacg gcgggcggct ggcgtggtgc
     2101 tggagatgat ccgggaaggg aagattgccg gtcgggcagt ccttattgct ggccagccgg
     2161 gcacggggaa gacggccatc gccatgggca tggcgcaggc cctgggccct gacacgccat
     2221 tcacagccat cgccggcagt gaaatcttct ccctggagat gagcaagacc gaggcgctga
     2281 cgcaggcctt ccggcggtcc atcggcgttc gcatcaagga ggagacggag atcatcgaag
     2341 gggaggtggt ggagatccag attgatcgac cagcaacagg gacgggctcc aaggtgggca
     2401 aactgaccct caagaccaca gagatggaga ccatctacga cctgggcacc aagatgattg
     2461 agtccctgac caaggacaag gtccaggccg gggacgtgat caccatcgac aaggcgacgg
     2521 gcaagatctc caagctgggc cgctccttca cacgcgcccg cgactacgac gctatgggct
     2581 cccagaccaa gttcgtgcag tgcccagatg gggagctcca gaaacgcaag gaggtggtgc
     2641 acaccgtgtc cctgcacgag atcgacgtca tcaactctcg cacccagggc ttcctggcgc
     2701 tcttctcagg tgacacaggg gagatcaagt cagaagtccg tgagcagatc aatgccaagg
     2761 tggctgagtg gcgcgaggag ggcaaggcgg agatcatccc tggagtgctg ttcatcgacg
     2821 aggtccacat gctggacatc gagagcttct ccttcctcaa ccgggccctg gagagtgaca
     2881 tggcgcctgt cctgatcatg gccaccaacc gtggcatcac gcgaatccgg ggcaccagct
     2941 accagagccc tcacggcatc cccatagacc tgctggaccg gctgcttatc gtctccacca
     3001 ccccctacag cgagaaagac acgaagcaga tcctccgcat ccggtgcgag gaagaagatg
     3061 tggagatgag tgaggacgcc tacacggtgc tgacccgcat cgggctggag acgtcactgc
     3121 gctacgccat ccagctcatc acagctgcca gcttggtgtg ccggaaacgc aagggtacag
     3181 aagtgcaggt ggatgacatc aagcgggtct actcactctt cctggacgag tcccgctcca
     3241 cgcagtacat gaaggagtac caggacgcct tcctcttcaa cgaactcaaa ggcgagacca
     3301 tggacacctc ctaagcttgc ggccgcactc gagcaccacc accaccacca ctgagatccg
     3361 gctgctaaca aagcccgaaa ggaagctgag ttggctgctg ccaccgctga gcaataacta
     3421 gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa aggaggaact
     3481 atatccggat tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     3541 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
     3601 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     3661 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg
     3721 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     3781 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
     3841 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     3901 agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttagg
     3961 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     4021 aaatatgtat ccgctcatga attaattctt agaaaaactc atcgagcatc aaatgaaact
     4081 gcaatttatt catatcagga ttatcaatac catatttttg aaaaagccgt ttctgtaatg
     4141 aaggagaaaa ctcaccgagg cagttccata ggatggcaag atcctggtat cggtctgcga
     4201 ttccgactcg tccaacatca atacaaccta ttaatttccc ctcgtcaaaa ataaggttat
     4261 caagtgagaa atcaccatga gtgacgactg aatccggtga gaatggcaaa agtttatgca
     4321 tttctttcca gacttgttca acaggccagc cattacgctc gtcatcaaaa tcactcgcat
     4381 caaccaaacc gttattcatt cgtgattgcg cctgagcgag acgaaatacg cgatcgctgt
     4441 taaaaggaca attacaaaca ggaatcgaat gcaaccggcg caggaacact gccagcgcat
     4501 caacaatatt ttcacctgaa tcaggatatt cttctaatac ctggaatgct gttttcccgg
     4561 ggatcgcagt ggtgagtaac catgcatcat caggagtacg gataaaatgc ttgatggtcg
     4621 gaagaggcat aaattccgtc agccagttta gtctgaccat ctcatctgta acatcattgg
     4681 caacgctacc tttgccatgt ttcagaaaca actctggcgc atcgggcttc ccatacaatc
     4741 gatagattgt cgcacctgat tgcccgacat tatcgcgagc ccatttatac ccatataaat
     4801 cagcatccat gttggaattt aatcgcggcc tagagcaaga cgtttcccgt tgaatatggc
     4861 tcataacacc ccttgtatta ctgtttatgt aagcagacag ttttattgtt catgaccaaa
     4921 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     4981 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     5041 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     5101 ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac
     5161 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     5221 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     5281 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     5341 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     5401 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     5461 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     5521 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     5581 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     5641 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     5701 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
     5761 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcaa tggtgcactc
     5821 tcagtacaat ctgctctgat gccgcatagt taagccagta tacactccgc tatcgctacg
     5881 tgactgggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     5941 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     6001 tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc
     6061 gtggtcgtga agcgattcac agatgtctgc ctgttcatcc gcgtccagct cgttgagttt
     6121 ctccagaagc gttaatgtct ggcttctgat aaagcgggcc atgttaaggg cggttttttc
     6181 ctgtttggtc actgatgcct ccgtgtaagg gggatttctg ttcatggggg taatgatacc
     6241 gatgaaacga gagaggatgc tcacgatacg ggttactgat gatgaacatg cccggttact
     6301 ggaacgttgt gagggtaaac aactggcggt atggatgcgg cgggaccaga gaaaaatcac
     6361 tcagggtcaa tgccagcgct tcgttaatac agatgtaggt gttccacagg gtagccagca
     6421 gcatcctgcg atgcagatcc ggaacataat ggtgcagggc gctgacttcc gcgtttccag
     6481 actttacgaa acacggaaac cgaagaccat tcatgttgtt gctcaggtcg cagacgtttt
     6541 gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct aaccagtaag
     6601 gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc acgatcatgc gcacccgtgg
     6661 ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa cgtttggtgg cgggaccagt
     6721 gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc gcaagcgaca ggccgatcat
     6781 cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc cagagcgctg ccggcacctg
     6841 tcctacgagt tgcatgataa agaagacagt cataagtgcg gcgacgatag tcatgccccg
     6901 cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc gagatcccgg
     6961 tgcctaatga gtgagctaac ttacattaat tgcgttgcgc
//
