LOCUS       sequence-457151-        8374 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express N-terminal three tandem FLAG epitope tags, followed by an 
            enterokinase cleavage site and human DGKD gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8374)
  AUTHORS   Murakami C, Hoshino F, Sakai H, Hayashi Y, Yamashita A, Sakane F
  TITLE     Diacylglycerol kinase delta and sphingomyelin synthase-related 
            protein functionally interact via their sterile alpha motif domains.
  JOURNAL   J Biol Chem. 2020 Mar 6;295(10):2932-2947. doi: 
            10.1074/jbc.RA119.012369. Epub 2020 Jan 24.
  PUBMED    31980461
REFERENCE   2  (bases 1 to 8374)
  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"; doi: 
            "10.1074/jbc.RA119.012369"; journalName: "J Biol Chem"; date: 
            "2020-03-6- 6"; volume: "295"; issue: "10"; pages: "2932-2947"
COMMENT     Sequence Label: 3xFLAG-DGK-delta-2
FEATURES             Location/Qualifiers
     source          1..8374
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    2661..3283
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2860..2879)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(3307..3327)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3312..3641
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3492..3627
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(3679..3698)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3680..3698)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3712..3728)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3712..3728)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3725..3747)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3736..3752
                     /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(3760..3766,3767..3784,3785..3790))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3805..3826
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3943..3960)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4114..4702)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4194..4213)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4873..5664,5665..5733))
                     /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     5496..5515
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5734..5838)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5865..6320
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5952..5971)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6162..6183
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6446..6468
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6460..6477
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6461..6477
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        6638..7017
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7018..7221
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7172..7192
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             7251..7316
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
ORIGIN
        1 actaaacccc gcccaggtct tcgacctcat gaacggaggc ccacacctcg gcttacggtt
       61 attccagaag tttgacacat tccggattct ggtttgtggc ggggatggaa gtgttggctg
      121 ggtcctctcc gaaatcgaca gcctcaacct tcataaacag tgtcagctgg gagtgctgcc
      181 gctcggcaca gggaacgact tggcccgagt actgggctgg ggctcagcct gcgatgacga
      241 cacccagctc ccccagatct tggagaagtt ggagagagcc agcaccaaga tgctggacag
      301 gtggagcgtc atggcatacg aggccaagct cccccggcag gcctcctcct ctaccgtcac
      361 cgaagacttc agcgaggatt ccgaggtaca gcagattctc ttctatgaag actcggttgc
      421 agcccacctt tctaaaatcc tcacctcgga ccagcactcg gtggtcatct cctcggccaa
      481 agtgctctgt gagacggtga aggacttcgt ggcacgggtg gggaaggcct atgagaagac
      541 gaccgagagc tcggaggagt cagaggtcat ggccaagaag tgctctgtcc tgaaagagaa
      601 gctggattcc cttctcaaga ccttggacga tgagtcccag gcctcgtcct ctctgcccaa
      661 cccgcccccc accattgccg aggaggctga agatggagat gggtcgggca gcatctgcgg
      721 ttccaccgga gaccgcttgg tggcatcagc ttgcccggcc cggccgcaga tattccggcc
      781 tcgagaacag ctcatgctga gagccaacag cctgaagaaa gcaattcgtc agatcataga
      841 acacacagaa aaagctgtcg atgagcagaa tgcccagacc caggagcagg agggcttcgt
      901 cctgggcctc tctgagtcag aggagaagat ggaccacaga gtgtgcccac cactgtccca
      961 cagcgagagc ttcggggtcc ccaaggggag gagccagcgc aaagtgtcga aatctccgtg
     1021 tgaaaagctg atcagcaaag ggagtctgtc cctaggcagt tctgcttccc ttccgcccca
     1081 gccgggaagc cgggacggcc tgcctgcgct caacaccaag atcctgtacc caaatgtccg
     1141 ggctggaatg tctggttcct tacccggtgg ctcagtcatc agtcgcctgt taattaatgc
     1201 tgatcccttc aactctgaac cagaaaccct agagtattac acggagaaat gtgtcatgaa
     1261 caactatttt ggcattggcc tggatgcgaa gatatccctg gactttaaca acaagcgcga
     1321 tgagcaccca gagaagtgca ggagccgaac caagaacatg atgtggtatg gagttcttgg
     1381 aaccaaagag ttgctgcaca gaacctacaa gaacctggag caaaaggtct tgctggagtg
     1441 tgacgggcga cccatcccac tccccagtct tcagggaatt gctgtcctta acattcccag
     1501 ctatgccgga ggaaccaact tctggggggg taccaaggaa gatgatactt tcgcagctcc
     1561 atcattcgat gacaagattc tggaggtggt cgccgtgttc ggcagcatgc agatggccgt
     1621 ctctcgagtc atcaggctac agcatcatcg gatcgcccag tgtcgcacgg tgaagatctc
     1681 catccttggg gatgagggcg tgcctgtgca ggtggacgga gaggcctggg tccagccgcc
     1741 agggtacatt cggattgtcc acaagaaccg ggcacagaca ctgaccagag acagggcatt
     1801 tgagagcacc ctgaagtcct gggaagacaa gcagaagtgc gagctgcccc gccctccatc
     1861 ctgttccctg cacccggaga tgctgtccga ggaggaggcc acccagatgg accagtttgg
     1921 gcaggcagca ggggtcctca ttcacagtat ccgagaaata gctcagtctc accgggacat
     1981 ggagcaggaa ctggcccacg ccgtcaatgc cagctccaag tccatggacc gtgtgtatgg
     2041 caagcccaga accacagagg ggctcaactg cagcttcgtc ctggaaatgg tgaataactt
     2101 cagagctctg cgcagtgaga cggagctgct gctgtctggg aagatggccc tgcagctgga
     2161 tccgcctcag aaggagcagc tggggagtgc tcttgccgag atggaccgac agctcaggag
     2221 gctggcagac accccgtggc tctgccagtc cgcagagccc ggcgacgaag agagtgtgat
     2281 gctggatctt gccaagcgca gtcgcagtgg taaattccgc ctcgtgacca agtttaaaaa
     2341 ggagaaaaac aacaagaaca aagaagctca cagtagcctg ggagccccgg ttcacctctg
     2401 ggggacagag gaggttgctg cctggctgga gcacctcagt ctctgtgagt ataaggacat
     2461 cttcacacgg cacgacatcc ggggctctga gctcctgcac ctggagcgga gggacctcaa
     2521 ggacctgggc gtgaccaagg tgggccacat gaagaggatc ctgtgtggca tcaaggagct
     2581 gagccgcagc gcccccgccg tcgaggccta gaattcatcg atagatctga tatcggtacc
     2641 agtcgactct agaggatccc gggtggcatc cctgtgaccc ctccccagtg cctctcctgg
     2701 ccctggaagt tgccactcca gtgcccacca gccttgtcct aataaaatta agttgcatca
     2761 ttttgtctga ctaggtgtcc ttctataata ttatggggtg gaggggggtg gtatggagca
     2821 aggggcaagt tgggaagaca acctgtaggg cctgcggggt ctattgggaa ccaagctgga
     2881 gtgcagtggc acaatcttgg ctcactgcaa tctccgcctc ctgggttcaa gcgattctcc
     2941 tgcctcagcc tcccgagttg ttgggattcc aggcatgcat gaccaggctc agctaatttt
     3001 tgtttttttg gtagagacgg ggtttcacca tattggccag gctggtctcc aactcctaat
     3061 ctcaggtgat ctacccacct tggcctccca aattgctggg attacaggcg tgaaccactg
     3121 ctcccttccc tgtccttctg attttaaaat aactatacca gcaggaggac gtccagacac
     3181 agcataggct acctggccat gcccaaccgg tgggacattt gagttgcttg cttggcactg
     3241 tcctctcatg cgttgggtcc actcagtaga tgcctgttga attgggtacg cggccagctt
     3301 ggctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     3361 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     3421 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     3481 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     3541 gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag ctattccaga
     3601 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcctc gaggaactga
     3661 aaaaccagaa agttaattcc ctatagtgag tcgtattaaa ttcgtaatca tgtcatagct
     3721 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     3781 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     3841 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     3901 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     3961 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     4021 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     4081 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     4141 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     4201 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     4261 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     4321 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     4381 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     4441 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     4501 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     4561 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     4621 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     4681 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     4741 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     4801 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     4861 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     4921 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     4981 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     5041 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     5101 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     5161 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     5221 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     5281 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     5341 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     5401 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     5461 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     5521 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     5581 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     5641 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     5701 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     5761 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     5821 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgcgc cctgtagcgg
     5881 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     5941 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     6001 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     6061 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     6121 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     6181 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     6241 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa
     6301 aatattaacg cttacaattt ccattcgcca ttcaggctgc gcaactgttg ggaagggcga
     6361 tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga
     6421 ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgcc
     6481 aagctgatct atacattgaa tcaatattgg caattagcca tattagtcat tggttatata
     6541 gcataaatca atattggcta ttggccattg catacgttgt atctatatca taatatgtac
     6601 atttatattg gctcatgtcc aatatgaccg ccatgttgac attgattatt gactagttat
     6661 taatagtaat caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca
     6721 taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca
     6781 ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg
     6841 gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtccg
     6901 ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc
     6961 ttacgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg
     7021 atgcggtttt ggcagtacac caatgggcgt ggatagcggt ttgactcacg gggatttcca
     7081 agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt
     7141 ccaaaatgtc gtaataaccc cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg
     7201 gaggtctata taagcagagc tcgtttagtg aaccgtcaga attaaccatg gactacaaag
     7261 accatgacgg tgattataaa gatcatgaca tcgattacaa ggatgacgat gacaagcttg
     7321 cggccgcgaa ttccggcagc atggcggcgg cggcgggcgc ccctccgccg ggtcccccgc
     7381 aaccgcctcc gccgccgccg cccgaggagt cgtccgacag cgagcccgag gcggagcccg
     7441 gctccccaca gaagctcatc cgcaaggtgt ccacgtcggg tcagatccga cagaagacca
     7501 tcatcaaaga ggggatgctg accaaacaga acaattcatt ccagcgatca aaaaggagat
     7561 actttaagct tcgagggcga acgctttact atgccaaaac ggcaaagtca atcatatttg
     7621 atgaggtgga tctgacagat gccagcgtag ctgaatccag taccaaaaac gtcaacaaca
     7681 gttttacggt cataactcca tgcaggaagc tcatcttgtg tgctgataac agaaaagaaa
     7741 tggaagattg gattgcagca ttaaagactg tgcagaacag ggagcacttt gagcccaccc
     7801 agtacagcat ggaccacttc tcagggatgc acaattggta cgcctgttcc cacgcgaggc
     7861 cgacctactg caatgtgtgc cgtgaggctc tgtctggggt cacgtcgcac gggctgtcct
     7921 gcgaggtgtg caaatttaag gcccacaagc gctgtgctgt gcgtgcaacc aataactgca
     7981 agtggaccac actggcctcg atcgggaagg acatcattga agatgcagat gggattgcaa
     8041 tgccccacca gtggttggaa ggaaacctac ctgtgagcgc caagtgcact gtgtgcgaca
     8101 agacctgtgg cagtgtgctg cgcctgcagg actggcgctg cctctggtgc aaggccatgg
     8161 ttcacacatc gtgtaaagaa tccttgctga ccaagtgccc acttggcctg tgcaaagtgt
     8221 cagtcatccc acccacggct ctcaacagca tcgactccga tgggttctgg aaggccagct
     8281 gtcctccttc ttgcacaagc ccactgttgg tcttcgtcaa ttcaaaaagt ggggacaacc
     8341 agggtgtgaa gttcctcaga agattcaaac agct
//
