LOCUS       sequence-473492-        7449 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  transient overexpression of FBXO21 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7449)
  AUTHORS   Donato V, Bonora M, Simoneschi D, Sartini D, Kudo Y, Saraf A, 
            Florens L, Washburn MP, Stadtfeld M, Pinton P, Pagano M
  TITLE     The TDH-GCN5L1-Fbxo15-KBP axis limits mitochondrial biogenesis in 
            mouse embryonic stem cells.
  JOURNAL   Nat Cell Biol. 2017 Apr;19(4):341-351. doi: 10.1038/ncb3491. Epub 
            2017 Mar 20.
  PUBMED    28319092
REFERENCE   2  (bases 1 to 7449)
  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: "10.1038/ncb3491"; 
            journalName: "Nat Cell Biol"; date: "2017-04"; volume: "19"; issue: 
            "4"; pages: "341-351"
COMMENT     Sequence Label: pcDNA3 Step-FLAG FBXO21
FEATURES             Location/Qualifiers
     source          1..7449
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     3..22
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(241..345)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(420..439)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        611..990
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        991..1194
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1145..1165
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1239..1258
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1239..1257
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1323..1332
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1332..1355
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1371..1394
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1431..1454
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     promoter        complement(3378..3396)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3379..3396)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3416..3433)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3422..3646
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3692..4120
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3779..3798)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3989..4010
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4129..4149)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4134..4463
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4314..4449
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4376..4395
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4530..5324
                     /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(4584..4603)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5194..5213
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5498..5619
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5535..5554)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5589..5608
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5668..5684)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5668..5684)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5681..5703)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5692..5708
                     /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(5716..5722,5723..5740,5741..5746))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5761..5782
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5899..5916)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6070..6658)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6150..6169)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6829..7449,1..171,172..240))
                     /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"
ORIGIN
        1 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
       61 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
      121 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
      181 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
      241 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
      301 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      361 agtgccacct gacgtcgacg gatcgggaga tctcccgatc ccctatggtg cactctcagt
      421 acaatctgct ctgatgccgc atagttaagc cagtatctgc tccctgcttg tgtgttggag
      481 gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa ggcaaggctt gaccgacaat
      541 tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc ttcgcgatgt acgggccaga
      601 tatacgcgtt gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta
      661 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
      721 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
      781 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
      841 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
      901 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
      961 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
     1021 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     1081 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
     1141 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg
     1201 ctaactagag aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag
     1261 acccaagctt ggtaccgagc tcggatccac tagtaacggc cgccagtgtg ctggaattcg
     1321 ccgccaccat ggattataaa gatgatgatg ataaagggtc ggccgccagc tggagccacc
     1381 ctcagttcga gaagggagga ggaagcggcg gaggcagcgg aggaggaagc tggagccacc
     1441 cgcagttcga gaaaggagct agcatggcgg cggcagcagt cgacagcgcg atggaggtgg
     1501 tgccggcgct ggcggaggag gccgcgccgg aggtagcggg cctcagctgc ctcgtcaacc
     1561 tgccgggtga ggtgctggag tacatcctgt gctgcggctc gctgacggcc gccgacatcg
     1621 gccgtgtctc cagcacctgc cggcggctgc gcgagctgtg ccagagcagc gggaaggtgt
     1681 ggaaggagca gttccgggtg aggtggcctt cccttatgaa acactacagc cccaccgact
     1741 acgtcaattg gttggaagag tataaagttc ggcaaaaagc tgggttagaa gcgcggaaga
     1801 ttgtagcctc gttctcaaag aggttctttt cagagcacgt tccttgtaat ggcttcagtg
     1861 acattgagaa ccttgaagga ccagagattt tttttgagga tgaactggtg tgtatcctaa
     1921 atatggaagg aagaaaagct ttgacctgga aatactacgc aaaaaaaatt ctttactacc
     1981 tgcggcaaca gaagatctta aataatctta aggcctttct tcagcagcca gatgactatg
     2041 agtcgtatct tgaaggtgct gtacatattg accagtactg caatcctctc tccgacatca
     2101 gcctcaaaga catccaggcc caaattgaca gcatcgtgga gcttgtttgc aaaacccttc
     2161 ggggcataaa cagtcgccac cccagcttgg ccttcaaggc aggtgaatca tccatgataa
     2221 tggaaataga actccagagc caggtgctgg atgccatgaa ctatgtcctt tacgaccaac
     2281 tgaagttcaa ggggaatcga atggattact ataatgccct caacttatat atgcatcagg
     2341 ttttgattcg cagaacagga atcccaatca gcatgtctct gctctatttg acaattgctc
     2401 ggcagttggg agtcccactg gagcctgtca acttcccaag tcacttctta ttaaggtggt
     2461 gccaaggcgc agaaggggcg accctggaca tctttgacta catctacata gatgcttttg
     2521 ggaaaggcaa gcagctgaca gtgaaagaat gcgagtactt gatcggccag cacgtgactg
     2581 cagcactgta tggggtggtc aatgtcaaga aggtgttaca gagaatggtg ggaaacctgt
     2641 taagcctggg gaagcgggaa ggcatcgacc agtcatacca gctcctgaga gactcgctgg
     2701 atctctatct ggcaatgtac ccggaccagg tgcagcttct cctcctccaa gccaggcttt
     2761 acttccacct gggaatctgg ccagagaagt ctttctgtct tgttttgaag gtgcttgaca
     2821 tcctccagca catccaaacc ctagacccgg ggcagcacgg ggcggtgggc tacctggtgc
     2881 agcacactct agagcacatt gagcgcaaaa aggaggaggt gggcgtagag gtgaagctgc
     2941 gctccgatga gaagcacaga gatgtctgct actccatcgg gctcattatg aagcataaga
     3001 ggtatggcta taactgtgtg atctacggct gggaccccac ctgcatgatg ggacacgagt
     3061 ggatccggaa catgaacgtc cacagcctgc cgcacggcca ccaccagcct ttctataacg
     3121 tgctggtgga ggacggctcc tgtcgatacg cagcccaaga aaacttggaa tataacgtgg
     3181 agcctcaaga aatctcacac cctgacgtgg gacgctattt ctcagagttt actggcactc
     3241 actacatccc aaacgcagag ctggagatcc ggtatccaga ggatctggag tttgtctatg
     3301 aaacggtgca gaatatttac agtgcaaaga aagagaacat agatgagtaa ctcgagcatg
     3361 catctagagg gccctattct atagtgtcac ctaaatgcta gagctcgctg atcagcctcg
     3421 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc
     3481 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt
     3541 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat
     3601 tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa
     3661 agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg
     3721 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     3781 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     3841 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     3901 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     3961 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     4021 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     4081 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc
     4141 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     4201 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     4261 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     4321 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     4381 atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt
     4441 ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg
     4501 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt
     4561 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct
     4621 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga
     4681 ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg
     4741 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact
     4801 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg
     4861 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct
     4921 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg
     4981 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt
     5041 tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg
     5101 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc
     5161 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag
     5221 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt
     5281 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt
     5341 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc
     5401 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
     5461 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa
     5521 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     5581 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac
     5641 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     5701 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     5761 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     5821 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     5881 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     5941 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     6001 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     6061 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     6121 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     6181 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     6241 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     6301 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     6361 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     6421 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     6481 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     6541 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     6601 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     6661 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     6721 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     6781 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     6841 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     6901 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     6961 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     7021 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     7081 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     7141 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     7201 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     7261 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     7321 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     7381 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     7441 gtcaatacg
//
