LOCUS       sequence-226677-        5976 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses HA-UBE2E1 C131A catalytic mutant in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5976)
  AUTHORS   Pasupala N, Morrow ME, Que LT, Malynn BA, Ma A, Wolberger C
  TITLE     OTUB1 non-catalytically stabilizes the E2 ubiquitin-conjugating 
            enzyme UBE2E1 by preventing its autoubiquitination.
  JOURNAL   J Biol Chem. 2018 Nov 23;293(47):18285-18295. doi: 
            10.1074/jbc.RA118.004677. Epub 2018 Oct 3.
  PUBMED    30282802
REFERENCE   2  (bases 1 to 5976)
  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.RA118.004677"; journalName: "J Biol Chem"; date: 
            "2018-11-23- 23"; volume: "293"; issue: "47"; pages: "18285-18295"
COMMENT     Sequence Label: HA-UBE2E1 C131A in pcDNA3.1
FEATURES             Location/Qualifiers
     source          1..5976
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             663..689
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     promoter        complement(1295..1313)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1296..1313)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1333..1350)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1339..1563
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1609..2037
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1696..1715)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1906..1927
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2046..2066)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2051..2380
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2231..2366
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2293..2312
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2447..3241
                     /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(2501..2520)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3111..3130
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3415..3536
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3452..3471)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3506..3525
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3585..3601)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3585..3601)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3598..3620)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3609..3625
                     /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(3633..3639,3640..3657,3658..3663))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3678..3699
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3816..3833)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3987..4575)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4067..4086)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4746..5537,5538..5606))
                     /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     5369..5388
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5607..5711)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5786..5805)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagcta
      661 tgtacccata cgatgttcca gattacgctt cggatgacga ttcgagggcc agcaccagct
      721 cctcctcatc ttcgtcttcc aaccagcaaa ccgagaaaga aacaaacacc cccaagaaga
      781 aggagagtaa agtcagcatg agcaaaaact ccaaactcct ctccaccagc gccaagagaa
      841 ttcagaagga gctggcggac atcactttag accctccacc taattgcagt gctggtccca
      901 aaggcgataa catctatgaa tggagatcaa ccattctagg gcctccagga tccgtgtatg
      961 agggtggtgt attctttctc gatatcactt ttacaccaga atatcccttc aagcctccaa
     1021 aggttacatt tcggacaaga atctatcatt gtaatattaa cagtcaaggt gttattgcct
     1081 tggacatatt gaaagataat tggagtccag cactaaccat ttctaaagtc ctcctttcta
     1141 tctgctcact tcttacagac tgtaatcctg ccgacccctt ggtgggaagt attgccactc
     1201 agtatatgac caacagagca gaacatgaca gaatggccag acagtggacc aagagatacg
     1261 ctacataatc gagcatgcat ctagagggcc ctattctata gtgtcaccta aatgctagag
     1321 ctcgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc
     1381 ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg
     1441 aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg
     1501 acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta
     1561 tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta
     1621 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     1681 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     1741 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     1801 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     1861 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     1921 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     1981 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat
     2041 tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag
     2101 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc
     2161 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct
     2221 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
     2281 actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa
     2341 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat
     2401 atccattttc ggatctgatc aagagacagg atgaggatcg tttcgcatga ttgaacaaga
     2461 tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc
     2521 acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc
     2581 ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc
     2641 gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac
     2701 tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc
     2761 tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac
     2821 gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg
     2881 tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct
     2941 cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt
     3001 cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg
     3061 attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac
     3121 ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg
     3181 tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg
     3241 agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat
     3301 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     3361 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
     3421 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     3481 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     3541 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     3601 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     3661 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     3721 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     3781 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     3841 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     3901 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     3961 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     4021 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     4081 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     4141 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     4201 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     4261 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     4321 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     4381 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     4441 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     4501 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     4561 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     4621 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     4681 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     4741 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     4801 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     4861 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     4921 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     4981 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     5041 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     5101 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     5161 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     5221 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     5281 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     5341 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     5401 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     5461 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     5521 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     5581 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     5641 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     5701 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatctc
     5761 ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
     5821 atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta
     5881 caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg
     5941 cgctgcttcg cgatgtacgg gccagatata cgcgtt
//
