LOCUS       sequence-246548-        5862 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses UBE2QL1 with C-terminal Strep-HA tag in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5862)
  AUTHORS   Koerver L, Papadopoulos C, Liu B, Kravic B, Rota G, Brecht L, 
            Veenendaal T, Polajnar M, Bluemke A, Ehrmann M, Klumperman J, 
            Jaattela M, Behrends C, Meyer H
  TITLE     The ubiquitin-conjugating enzyme UBE2QL1 coordinates lysophagy in 
            response to endolysosomal damage.
  JOURNAL   EMBO Rep. 2019 Aug 21:e48014. doi: 10.15252/embr.201948014.
  PUBMED    31432621
REFERENCE   2  (bases 1 to 5862)
  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"; journalName: "EMBO Rep. 
            2019 Aug 21:e48014. doi: 10.15252/embr.201948014."
COMMENT     Sequence Label: pcDNA5FRT/TO-UBE2QL1-Strep/HA
FEATURES             Location/Qualifiers
     source          1..5862
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        142..521
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        522..725
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     676..696
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    727..745
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    748..766
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     776..800
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    943..967
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1466..1490)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             1537..1560
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1597..1620
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             1651..1677
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             1651..1677
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(1721..1738)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1727..1951
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2084..2103)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2235..2282
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             2290..3312
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2290..3312
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    3442..3563
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3479..3498)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3533..3552
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3612..3628)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3612..3628)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3625..3647)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3636..3652
                     /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(3660..3666,3667..3684,3685..3690))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3705..3726
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3843..3860)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4014..4602)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4094..4113)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4773..5564,5565..5633))
                     /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     5396..5415
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5634..5738)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5813..5832)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct
       61 tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg
      121 tacgggccag atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta
      181 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
      241 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
      301 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
      361 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca
      421 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta
      481 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
      541 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
      601 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca
      661 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
      721 gagctctccc tatcagtgat agagatctcc ctatcagtga tagagatcgt cgacgagctc
      781 gtttagtgaa ccgtcagatc gcctggagac gccatccacg ctgttttgac ctccatagaa
      841 gacaccggga ccgatccagc ctccggactc tagcgtttaa acttaagctt ggtaccgagc
      901 tcggatccac tagtccagtg tggtggaatt ctgcagatat caacaagttt gtacaaaaaa
      961 gcaggcttct cgagcatgaa ggagctgcag gacatcgcgc gccttagcga ccgcttcatc
     1021 tccgtggagc tggtggacga gagcctgttc gactggaacg tgaagctgca ccaggtggac
     1081 aaggactcgg tgctgtggca ggacatgaag gagaccaaca ccgagttcat cctgctcaac
     1141 ctcaccttcc ccgacaactt ccccttctcg ccgcccttca tgcgggtgct cagcccgcgc
     1201 ctggagaacg gctacgtgct ggacggcggc gccatctgca tggagctgct cacgccgcgc
     1261 ggctggtcca gcgcctacac cgtggaggcc gtcatgcgcc agttcgcagc cagcctggtc
     1321 aagggccagg gacggatctg tagaaaagct ggcaaatcaa aaaagtcctt cagtcgcaag
     1381 gaagctgaag ctacctttaa gagtttggtg aagacgcatg aaaaatatgg ttgggtcacc
     1441 ccgcccgtgt ccgacggcgg aattcaccca gctttcttgt acaaagtggt tgatatcaag
     1501 gccgccgaca tcaccagcct gtacaagaaa gtggggtgga gccatcccca gttcgagaag
     1561 ggcggaggat ctggcggcgg aagcggcgga ggaagctggt cccaccctca gtttgagaaa
     1621 ggcaccgagc tgggcagcac catggccagc tacccctacg acgtgcccga ctacgcctga
     1681 tgactcgagt ctagagggcc cgtttaaacc cgctgatcag cctcgactgt gccttctagt
     1741 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact
     1801 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat
     1861 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc
     1921 aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc
     1981 tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     2041 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     2101 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     2161 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
     2221 ggttcacgta cctagaagtt cctattccga agttcctatt ctctagaaag tataggaact
     2281 tccttggcca aaaagcctga actcaccgcg acgtctgtcg agaagtttct gatcgaaaag
     2341 ttcgacagcg tctccgacct gatgcagctc tcggagggcg aagaatctcg tgctttcagc
     2401 ttcgatgtag gagggcgtgg atatgtcctg cgggtaaata gctgcgccga tggtttctac
     2461 aaagatcgtt atgtttatcg gcactttgca tcggccgcgc tcccgattcc ggaagtgctt
     2521 gacattgggg aattcagcga gagcctgacc tattgcatct cccgccgtgc acagggtgtc
     2581 acgttgcaag acctgcctga aaccgaactg cccgctgttc tgcagccggt cgcggaggcc
     2641 atggatgcga tcgctgcggc cgatcttagc cagacgagcg ggttcggccc attcggaccg
     2701 caaggaatcg gtcaatacac tacatggcgt gatttcatat gcgcgattgc tgatccccat
     2761 gtgtatcact ggcaaactgt gatggacgac accgtcagtg cgtccgtcgc gcaggctctc
     2821 gatgagctga tgctttgggc cgaggactgc cccgaagtcc ggcacctcgt gcacgcggat
     2881 ttcggctcca acaatgtcct gacggacaat ggccgcataa cagcggtcat tgactggagc
     2941 gaggcgatgt tcggggattc ccaatacgag gtcgccaaca tcttcttctg gaggccgtgg
     3001 ttggcttgta tggagcagca gacgcgctac ttcgagcgga ggcatccgga gcttgcagga
     3061 tcgccgcggc tccgggcgta tatgctccgc attggtcttg accaactcta tcagagcttg
     3121 gttgacggca atttcgatga tgcagcttgg gcgcagggtc gatgcgacgc aatcgtccga
     3181 tccggagccg ggactgtcgg gcgtacacaa atcgcccgca gaagcgcggc cgtctggacc
     3241 gatggctgtg tagaagtact cgccgatagt ggaaaccgac gccccagcac tcgtccgagg
     3301 gcaaaggaat agcacgtact acgagatttc gattccaccg ccgccttcta tgaaaggttg
     3361 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     3421 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     3481 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     3541 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     3601 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     3661 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     3721 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     3781 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     3841 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     3901 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     3961 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     4021 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     4081 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     4141 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     4201 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     4261 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     4321 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     4381 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     4441 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     4501 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     4561 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     4621 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     4681 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     4741 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     4801 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     4861 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     4921 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     4981 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     5041 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     5101 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     5161 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     5221 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     5281 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     5341 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     5401 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     5461 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     5521 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     5581 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     5641 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     5701 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     5761 cctgacgtcg acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct
     5821 gctctgatgc cgcatagtta agccagtatc tgctccctgc tt
//
