LOCUS       Exported                6351 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Mammalian expression of human SIRT5 with flag tag.
ACCESSION   .
VERSION     .
KEYWORDS    SIRT5 Flag
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6351)
  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 6351)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Mol Cell. 
            2003 Feb . 11(2):437-44."
FEATURES             Location/Qualifiers
     source          1..6351
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(63..80)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(234..822)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(314..333)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(993..1853)
                     /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     1616..1635
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1854..1958)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2033..2052)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2224..2603
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2604..2807
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2758..2778
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2852..2871
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2852..2870
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3873..3896
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(3931..3948)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3937..4161
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4207..4635
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4294..4313)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4504..4525
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4644..4664)
                     /label=pBABE 3
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4649..4978
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4829..4964
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4891..4910
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5045..5839
                     /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(5099..5118)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5709..5728
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6013..6134
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6050..6069)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6104..6123
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6183..6199)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6183..6199)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6196..6218)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6207..6223
                     /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(6231..6261)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6276..6297
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."