LOCUS       sequence-364363-        7414 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of human SIRT1 with flag tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7414)
  AUTHORS   North BJ, Marshall BL, Borra MT, Denu JM, Verdin E
  TITLE     The human Sir2 ortholog, SIRT2, is an NAD+-dependent tubulin 
            deacetylase.
  JOURNAL   Mol Cell. 2003 Feb . 11(2):437-44.
  PUBMED    12620231
REFERENCE   2  (bases 1 to 7414)
  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: "Mol Cell. 
            2003 Feb . 11(2):437-44."
COMMENT     Sequence Label: SIRT1 Flag
FEATURES             Location/Qualifiers
     source          1..7414
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    2..18
                     /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(26..32,33..50,51..56))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    71..92
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(209..226)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(380..968)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(460..479)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1139..1930,1931..1999))
                     /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     1762..1781
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2000..2104)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2179..2198)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2370..2749
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2750..2953
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2904..2924
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2998..3017
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2998..3016
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5082..5105
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(5140..5157)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5146..5370
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      5416..5844
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5503..5522)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5713..5734
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5853..5873)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5858..6187
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6038..6173
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6100..6119
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             6254..7048
                     /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(6308..6327)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6918..6937
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    7222..7343
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7259..7278)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7313..7332
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(7392..7408)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7392..7408)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(join(7405..7414,1..13))
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
ORIGIN
        1 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
       61 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
      121 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
      181 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
      241 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
      301 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
      361 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
      421 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
      481 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
      541 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
      601 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
      661 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
      721 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
      781 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
      841 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
      901 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
      961 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     1021 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     1081 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     1141 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     1201 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     1261 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     1321 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     1381 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     1441 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     1501 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     1561 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     1621 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     1681 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     1741 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     1801 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     1861 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     1921 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     1981 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     2041 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     2101 cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg atcgggagat ctcccgatcc
     2161 cctatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agtatctgct
     2221 ccctgcttgt gtgttggagg tcgctgagta gtgcgcgagc aaaatttaag ctacaacaag
     2281 gcaaggcttg accgacaatt gcatgaagaa tctgcttagg gttaggcgtt ttgcgctgct
     2341 tcgcgatgta cgggccagat atacgcgttg acattgatta ttgactagtt attaatagta
     2401 atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac
     2461 ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac
     2521 gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt
     2581 acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat
     2641 tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga
     2701 ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt
     2761 ttggcagtac atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca
     2821 ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg
     2881 tcgtaacaac tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta
     2941 tataagcaga gctctctggc taactagaga acccactgct tactggctta tcgaaattaa
     3001 tacgactcac tatagggaga cccaagctgg ctagcgttta aacttaagct tgcggccgca
     3061 tggcggacga ggcggcggcg gcaggcgggg agcaagaggc ccaggcgact gcggcggctg
     3121 gggaaggaga caatgggccg ggcctgcagg gcccatctcg ggagccaccg ctggccgaca
     3181 acttgtacga cgaagacgac gacgacgagg gcgaggagga ggaagaggcg gcggcggcgg
     3241 cgattgggta ccgagataac cttctgttcg gtgatgaaat tatcactaat ggttttcatt
     3301 cctgtgaaag tgatgaggag gatagagcct cacatgcaag ctctagtgac tggactccaa
     3361 ggccacggat aggtccatat acttttgttc agcaacatct tatgattggc acagatcctc
     3421 gaacaattct taaagattta ttgccggaaa caatacctcc acctgagttg gatgatatga
     3481 cactgtggca gattgttatt aatatccttt cagaaccacc aaaaaggaaa aaaagaaaag
     3541 atattaatac aattgaagat gctgtgaaat tactgcaaga gtgcaaaaaa attatagttc
     3601 taactggagc tggggtgtct gtttcatgtg gaatacctga cttcaggtca agggatggta
     3661 tttatgctcg ccttgctgta gacttcccag atcttccaga tcctcaagcg atgtttgata
     3721 ttgaatattt cagaaaagat ccaagaccat tcttcaagtt tgcaaaggaa atatatcctg
     3781 gacaattcca gccatctctc tgtcacaaat tcatagcctt gtcagataag gaaggaaaac
     3841 tacttcgcaa ctatacccag aacatagaca cgctggaaca ggttgcggga atccaaagga
     3901 taattcagtg tcatggttcc tttgcaacag catcttgcct gatttgtaaa tacaaagttg
     3961 actgtgaagc tgtacgagga gatattttta atcaggtagt tcctcgatgt cctaggtgcc
     4021 cagctgatga accgcttgct atcatgaaac cagagattgt gttttttggt gaaaatttac
     4081 cagaacagtt tcatagagcc atgaagtatg acaaagatga agttgacctc ctcattgtta
     4141 ttgggtcttc cctcaaagta agaccagtag cactaattcc aagttccata ccccatgaag
     4201 tgcctcagat attaattaat agagaacctt tgcctcatct gcattttgat gtagagcttc
     4261 ttggagactg tgatgtcata attaatgaat tgtgtcatag gttaggtggt gaatatgcca
     4321 aactttgctg taaccctgta aagctttcag aaattactga aaaacctcca cgaacacaaa
     4381 aagaattggc ttatttgtca gagttgccac ccacacctct tcatgtttca gaagactcaa
     4441 gttcaccaga aagaacttca ccaccagatt cttcagtgat tgtcacactt ttagaccaag
     4501 cagctaagag taatgatgat ttagatgtgt ctgaatcaaa aggttgtatg gaagaaaaac
     4561 cacaggaagt acaaacttct aggaatgttg aaagtattgc tgaacagatg gaaaatccgg
     4621 atttgaagaa tgttggttct agtactgggg agaaaaatga aagaacttca gtggctggaa
     4681 cagtgagaaa atgctggcct aatagagtgg caaaggagca gattagtagg cggcttgatg
     4741 gtaatcagta tctgtttttg ccaccaaatc gttacatttt ccatggcgct gaggtatatt
     4801 cagactctga agatgacgtc ttatcctcta gttcttgtgg cagtaacagt gatagtggga
     4861 catgccagag tccaagttta gaagaaccca tggaggatga aagtgaaatt gaagaattct
     4921 acaatggctt agaagatgag cctgatgttc cagagagagc tggaggagct ggatttggga
     4981 ctgatggaga tgatcaagag gcaattaatg aagctatatc tgtgaaacag gaagtaacag
     5041 acatgaacta tccatcaaac aaatcaagcg gccgcggagg agactacaag gacgacgatg
     5101 acaagtaatc tagagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt
     5161 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     5221 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     5281 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     5341 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct
     5401 ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     5461 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     5521 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     5581 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     5641 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     5701 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     5761 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     5821 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
     5881 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     5941 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     6001 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     6061 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     6121 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     6181 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcaag agacaggatg
     6241 aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt
     6301 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
     6361 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
     6421 cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
     6481 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
     6541 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
     6601 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
     6661 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
     6721 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
     6781 gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
     6841 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
     6901 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
     6961 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
     7021 ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa
     7081 gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg
     7141 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     7201 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     7261 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     7321 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     7381 gcgtaatcat ggtcatagct gtttcctgtg tgaa
//
