LOCUS       sequence-483738-        6724 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 6724)
  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 6724)
  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-DEST40-SIRT3
FEATURES             Location/Qualifiers
     source          1..6724
                     /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    911..935
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2172..2196)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             2222..2263
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2273..2290
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2313..2330)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2319..2543
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2589..3017
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2676..2695)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2886..2907
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3026..3046)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3031..3360
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3211..3346
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3273..3292
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3427..4221
                     /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(3481..3500)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4091..4110
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4397..4518
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4434..4453)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4488..4507
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4567..4583)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4567..4583)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4580..4602)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4591..4607
                     /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(4615..4621,4622..4639,4640..4645))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4660..4681
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4798..4815)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4969..5557)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5049..5068)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5728..6519,6520..6588))
                     /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     6351..6370
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6589..6693)
                     /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 taagctatca acaagtttgt acaaaaaagc aggctccgcg gccgccccct tcaccatggc
      961 gttctggggt tggcgcgccg cggcagccct ccggctgtgg ggccgggtag ttgaacgggt
     1021 cgaggccggg ggaggcgtgg ggccgtttca ggcctgcggc tgtcggctgg tgcttggcgg
     1081 cagggacgat gtgagtgcgg ggctgagagg cagccatggg gcccgcggtg agcccttgga
     1141 cccggcgcgc cccttgcaga ggcctcccag acccgaggtg cccagggcat tccggaggca
     1201 gccgagggca gcagctccca gtttcttctt ttcgagtatt aaaggtggaa gaaggtccat
     1261 atctttttct gtgggtgctt caagtgttgt tggaagtgga ggcagcagtg acaaggggaa
     1321 gctttccctg caggatgtag ctgagctgat tcgggccaga gcctgccaga gggtggtggt
     1381 catggtgggg gccggcatca gcacacccag tggcattcca gacttcagat cgccggggag
     1441 tggcctgtac agcaacctcc agcagtacga tctcccgtac cccgaggcca tttttgaact
     1501 cccattcttc tttcacaacc ccaagccctt tttcactttg gccaaggagc tgtaccctgg
     1561 aaactacaag cccaacgtca ctcactactt tctccggctg cttcatgaca aggggctgct
     1621 tctgcggctc tacacgcaga acatcgatgg gcttgagaga gtgtcgggca tccctgcctc
     1681 aaagctggtt gaagctcatg gaacctttgc ctctgccacc tgcacagtct gccaaagacc
     1741 cttcccaggg gaggacattc gggctgacgt gatggcagac agggttcccc gctgcccggt
     1801 ctgcaccggc gttgtgaagc ccgacattgt gttctttggg gagccgctgc cccagaggtt
     1861 cttgctgcat gtggttgatt tccccatggc agatctgctg ctcatccttg ggacctccct
     1921 ggaggtggag ccttttgcca gcttgaccga ggccgtgcgg agctcagttc cccgactgct
     1981 catcaaccgg gacttggtgg ggcccttggc ttggcatcct cgcagcaggg acgtggccca
     2041 gctgggggac gtggttcacg gcgtggaaag cctagtggag cttctgggct ggacagaaga
     2101 gatgcgggac cttgtgcagc gggaaactgg gaagcttgat ggaccagaca aatttaaggg
     2161 tgggcgcgcc gacccagctt tcttgtacaa agtggttgat ctagagggcc cgcggttcga
     2221 aggtaagcct atccctaacc ctctcctcgg tctcgattct acgcgtaccg gtcatcatca
     2281 ccatcaccat tgagtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc
     2341 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     2401 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct
     2461 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc
     2521 tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg
     2581 gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     2641 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt
     2701 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     2761 ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg
     2821 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     2881 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt
     2941 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
     3001 aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca
     3061 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt
     3121 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     3181 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
     3241 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct
     3301 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa
     3361 aagctcccgg gagcttgtat atccattttc ggatctgatc aagagacagg atgaggatcg
     3421 tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg
     3481 ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg
     3541 ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat
     3601 gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca
     3661 gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg
     3721 gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat
     3781 gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa
     3841 catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg
     3901 gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg
     3961 cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg
     4021 gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat
     4081 caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac
     4141 cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc
     4201 cttcttgacg agttcttctg agcgggactc tggggttcgc gaaatgaccg accaagcgac
     4261 gcccaacctg ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
     4321 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
     4381 gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     4441 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     4501 actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta
     4561 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     4621 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     4681 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     4741 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     4801 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     4861 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     4921 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     4981 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     5041 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     5101 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     5161 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     5221 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     5281 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     5341 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     5401 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     5461 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     5521 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     5581 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     5641 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     5701 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     5761 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     5821 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     5881 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     5941 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     6001 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     6061 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     6121 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     6181 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     6241 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     6301 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     6361 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     6421 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     6481 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     6541 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     6601 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     6661 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     6721 cgtc
//
