LOCUS       sequence-483743-        7365 bp    DNA     circular SYN 18-AUG-2025
DEFINITION  Expression in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7365)
  AUTHORS   Michishita E, Park JY, Burneskis JM, Barrett JC, Horikawa I
  TITLE     Evolutionarily conserved and nonconserved cellular localizations and
            functions of human SIRT proteins.
  JOURNAL   Mol Biol Cell. 2005 Oct;16(10):4623-35. doi: 
            10.1091/mbc.e05-01-0033. Epub 2005 Aug 3.
  PUBMED    16079181
REFERENCE   2  (bases 1 to 7365)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 18, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1091/mbc.e05-01-0033"; journalName: "Mol Biol Cell"; date: 
            "2005-10"; volume: "16"; issue: "10"; pages: "4623-35"
COMMENT     Sequence Label: pcDNA-DEST47-SIRT7
FEATURES             Location/Qualifiers
     source          1..7365
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    914..938
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2178..2202)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             join(2213..2215,2216..2218,2219..2932)
                     /codon_start=1
                     /product="Cycle 3 GFP (Crameri et al., 1996)"
                     /label=Cycle 3 GFP
                     /note="mammalian codon-optimized"
                     /translation="MASKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITHGMDELYK"
     primer_bind     complement(2246..2274)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     2869..2889
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     complement(2954..2971)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2960..3184
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3230..3658
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3317..3336)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3527..3548
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3667..3687)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3672..4001
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3852..3987
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3914..3933
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4068..4862
                     /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(4122..4141)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4732..4751
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5038..5159
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5075..5094)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5129..5148
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5208..5224)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5208..5224)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5221..5243)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5232..5248
                     /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(5256..5262,5263..5280,5281..5286))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5301..5322
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5439..5456)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5610..6198)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5690..5709)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6369..7160,7161..7229))
                     /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     6992..7011
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7230..7334)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagt
      901 taagcttgat caaacaagtt tgtacaaaaa agcaggctcc gcggccgccc ccttcaccat
      961 ggcagccggg ggtctgagcc gctccgagcg caaagcggcg gagcgggtcc ggaggttgcg
     1021 ggaggagcag cagagggagc gcctccgcca ggtgtcgcgc atcctgagga aggcggcggc
     1081 ggagcgcagc gccgaggagg gccggctgct ggccgagagc gcggacctgg taacggagct
     1141 gcagggccgg agccggcggc gcgagggcct gaagcggcgg caggaggagg tgtgcgacga
     1201 cccggaggag ctgcggggga aggtccggga gctggccagc gccgtccgga acgccaaata
     1261 cttggtcgtc tacacaggcg cgggaatcag cacggcagcg tctatcccag actaccgggg
     1321 ccctaatgga gtgtggacac tgcttcagaa agggagaagc gttagtgctg ccgacctgag
     1381 cgaggccgag ccaaccctca cccacatgag catcacccgt ctgcatgagc agaagctggt
     1441 gcagcatgtg gtgtctcaga actgtgacgg gctccacctg aggagtgggc tgccgcgcac
     1501 ggccatctcc gagctccacg ggaacatgta cattgaagtc tgtacctcct gcgttcccaa
     1561 cagggagtac gtgcgggtgt tcgatgtgac ggagcgcact gccctccaca gacaccagac
     1621 aggccggacc tgccacaagt gtgggaccca gctgcgggac accattgtgc actttgggga
     1681 gagggggacg ttggggcagc ctctgaactg ggaagcggcg accgaggctg ccagcagagc
     1741 agacaccatc ctgtgtctag ggtccagcct gaaggttcta aagaagtacc cacgcctctg
     1801 gtgcatgacc aagcccccta gccggcggcc gaagctttac atcgtgaacc tgcagtggac
     1861 cccgaaggat gactgggctg ccctgaagct acatgggaag tgtgatgacg tcatgcggct
     1921 cctcatggcc gagctgggct tggagatccc cgcctatagc aggtggcagg atcccatttt
     1981 ctcactggcg actcccctgc gtgctggtga agaaggcagc cacagtcgga agtcgctgtg
     2041 cagaagcaga gaggaggccc cgcctgggga ccggggtgca ccgcttagct cggcccccat
     2101 cctagggggc tggtttggca ggggctgcac aaaacgcaca aaaaggaaga aagtgacgtt
     2161 taagggtggg cgcgccgacc cagctttctt gtacaaagtg gttcgatcta gaatggctag
     2221 caaaggagaa gaacttttca ctggagttgt cccaattctt gttgaattag atggtgatgt
     2281 taatgggcac aaattttctg tcagtggaga gggtgaaggt gatgctacat acggaaagct
     2341 tacccttaaa tttatttgca ctactggaaa actacctgtt ccatggccaa cacttgtcac
     2401 tactttctct tatggtgttc aatgcttttc ccgttatccg gatcatatga aacggcatga
     2461 ctttttcaag agtgccatgc ccgaaggtta tgtacaggaa cgcactatat ctttcaaaga
     2521 tgacgggaac tacaagacgc gtgctgaagt caagtttgaa ggtgataccc ttgttaatcg
     2581 tatcgagtta aaaggtattg attttaaaga agatggaaac attctcggac acaaactcga
     2641 gtacaactat aactcacaca atgtatacat cacggcagac aaacaaaaga atggaatcaa
     2701 agctaacttc aaaattcgcc acaacattga agatggatcc gttcaactag cagaccatta
     2761 tcaacaaaat actccaattg gcgatggccc tgtcctttta ccagacaacc attacctgtc
     2821 gacacaatct gccctttcga aagatcccaa cgaaaagcgt gaccacatgg tccttcttga
     2881 gtttgtaact gctgctggga ttacacatgg catggatgag ctctacaaat aatgaattaa
     2941 acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     3001 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     3061 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     3121 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct
     3181 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt
     3241 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     3301 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     3361 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     3421 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     3481 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     3541 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     3601 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa
     3661 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     3721 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     3781 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     3841 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     3901 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga
     3961 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta
     4021 tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag
     4081 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     4141 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     4201 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag
     4261 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     4321 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     4381 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     4441 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     4501 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     4561 tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg
     4621 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     4681 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     4741 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     4801 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     4861 gagcgggact ctggggttcg cgaaatgacc gaccaagcga cgcccaacct gccatcacga
     4921 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     4981 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     5041 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     5101 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     5161 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
     5221 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     5281 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     5341 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     5401 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     5461 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     5521 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     5581 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     5641 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     5701 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     5761 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     5821 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     5881 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     5941 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     6001 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     6061 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     6121 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     6181 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     6241 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     6301 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     6361 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     6421 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     6481 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     6541 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     6601 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     6661 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     6721 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     6781 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     6841 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     6901 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     6961 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     7021 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     7081 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     7141 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     7201 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     7261 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     7321 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
