LOCUS       sequence-139196-        7118 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7118)
  AUTHORS   Sowa ME, Bennett EJ, Gygi SP, Harper JW
  TITLE     Defining the human deubiquitinating enzyme interaction landscape.
  JOURNAL   Cell. 2009 Jul 23. 138(2):389-403.
  PUBMED    19615732
REFERENCE   2  (bases 1 to 7118)
  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: "Cell. 2009 
            Jul 23. 138(2):389-403."
COMMENT     Sequence Label: Flag-HA-UCHL3
FEATURES             Location/Qualifiers
     source          1..7118
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     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(join(993..1784,1785..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     2026..2044
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2082..2104)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2204..2223
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2417..2439
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2567..2586
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             2786..3302
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
     misc_feature    3366..3707
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             3774..4190
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     4120..4142
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     4158..4174
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      4203..4212
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4212..4235
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             4248..4274
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    4290..4314
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(5005..5029)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    5042..5609
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    5059..5611
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(5203..5220)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     5430..5449
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             5623..6222
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(5623..5642)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6119..6139
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             6298..6812
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    6974..6990
                     /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(6998..7004,7005..7022,7023..7028))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7043..7064
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
       61 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
      121 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
      181 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
      241 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
      301 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
      361 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
      421 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
      481 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
      541 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
      601 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
      661 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
      721 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
      781 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
      841 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
      901 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
      961 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     1021 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     1081 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     1141 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     1201 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     1261 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     1321 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     1381 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     1441 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     1501 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     1561 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     1621 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     1681 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     1741 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     1801 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     1861 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     1921 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     1981 cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg
     2041 aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc
     2101 ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc
     2161 gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact atgcggcatc agagcagatt
     2221 gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
     2281 cgcatcaggc gccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg
     2341 gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg
     2401 gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggcgcaagg aatggtgcat
     2461 gcaaggagat ggcgcccaac agtcccccgg ccacggggcc tgccaccata cccacgccga
     2521 aacaagcgct catgagcccg aagtggcgag cccgatcttc cccatcggtg atgtcggcga
     2581 tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt
     2641 agaggcgatt agtccaattt gttaaagaca ggatatcagt ggtccaggct ctagttttga
     2701 ctcaacaata tcaccagctg aagcctatag agtacgagcc atagataaaa taaaagattt
     2761 tatttagtct ccagaaaaag gggggaatga aagaccccac ctgtaggttt ggcaagctag
     2821 cttaagtaac gccattttgc aaggcatgga aaatacataa ctgagaatag agaagttcag
     2881 atcaaggtta ggaacagaga gacagcagaa tatgggccaa acaggatatc tgtggtaagc
     2941 agttcctgcc ccggctcagg gccaagaaca gatggtcccc agatgcggtc ccgccctcag
     3001 cagtttctag agaaccatca gatgtttcca gggtgcccca aggacctgaa atgaccctgt
     3061 gccttatttg aactaaccaa tcagttcgct tctcgcttct gttcgcgcgc ttctgctccc
     3121 cgagctcaat aaaagagccc acaacccctc actcggcgcg ccagtcctcc gatagactgc
     3181 gtcgcccggg tacccgtatt cccaataaag cctcttgctg tttgcatccg aatcgtggac
     3241 tcgctgatcc ttgggagggt ctcctcagat tgattgactg cccacctcgg gggtctttca
     3301 tttggaggtt ccaccgagat ttggagaccc ctgcccaggg accaccgacc cccccgccgg
     3361 gaggtaagct ggccagcggt cgtttcgtgt ctgtctctgt ctttgtgcgt gtttgtgccg
     3421 gcatctaatg tttgcgcctg cgtctgtact agttagctaa ctagctctgt atctggcgga
     3481 cccgtggtgg aactgacgag ttctgaacac ccggccgcaa ccctgggaga cgtcccaggg
     3541 actttggggg ccgtttttgt ggcccgacct gaggaaggga gtcgatgtgg aatccgaccc
     3601 cgtcaggata tgtggttctg gtaggagacg agaacctaaa acagttcccg cctccgtctg
     3661 aatttttgct ttcggtttgg aaccgaagcc gcgcgtcttg tctgctgcag cgctgcagca
     3721 tcgttctgtg ttgtctctgt ctgactgtgt ttctgtattt gtctgaaaat tagggccaga
     3781 ctgttaccac tcccttaagt ttgaccttag gtcactggaa agatgtcgag cggatcgctc
     3841 acaaccagtc ggtagatgtc aagaagagac gttgggttac cttctgctct gcagaatggc
     3901 caacctttaa cgtcggatgg ccgcgagacg gcacctttaa ccgagacctc atcacccagg
     3961 ttaagatcaa ggtcttttca cctggcccgc atggacaccc agaccaggtc ccctacatcg
     4021 tgacctggga agccttggct tttgaccccc ctccctgggt caagcccttt gtacacccta
     4081 agcctccgcc tcctcttcct ccatccgccc cgtctctccc ccttgaacct cctcgttcga
     4141 ccccgcctcg atcctccctt tatccagccc tcactccttc tctaggcgcc ggaattagat
     4201 ctgccaccat ggactacaag gatgacgatg acaagctcga tggaggatac ccatacgatg
     4261 ttccagatta cgctgctcga ggttatcaaa caagtttgta caaaaaagtt ggcatggagg
     4321 gtcaacgctg gctgccgctg gaggccaatc ccgaggtcac caaccagttt cttaaacaat
     4381 taggtctaca tcctaactgg caattcgttg atgtatatgg aatggatcct gaactcctta
     4441 gcatggtacc aagaccagtc tgtgcagtct tacttctctt tcctattaca gaaaagtatg
     4501 aagtattcag aacagaagag gaagaaaaaa taaaatctca gggacaagat gttacatcat
     4561 cagtatattt catgaagcaa acaatcagca atgcctgtgg aacaattgga ctgattcatg
     4621 ctattgcaaa caataaagac aagatgcact ttgaatctgg atcaaccttg aaaaaattcc
     4681 tggaggaatc tgtgtcaatg agccctgaag aacgagccag atacctggag aactatgatg
     4741 ccatccgagt tactcatgag accagtgccc atgaaggtca gactgaggca ccaagtatag
     4801 atgagaaagt agatcttcat tttattgcat tagttcatgt agatgggcat ctctatgaat
     4861 tagatgggcg gaagccattt ccaattaacc atggtgaaac tagtgatgaa actttattag
     4921 aggatgccat agaagtttgc aagaagttta tggagcgcga ccctgatgaa ctaagattta
     4981 atgcgattgc tctttctgca gcatacccaa ctttcttgta caaagtggtt cgataacgaa
     5041 ttccgccccc cccccctaac gttactggcc gaagccgctt ggaataaggc cggtgtgcgt
     5101 ttgtctatat gttattttcc accatattgc cgtcttttgg caatgtgagg gcccggaaac
     5161 ctggccctgt cttcttgacg agcattccta ggggtctttc ccctctcgcc aaaggaatgc
     5221 aaggtctgtt gaatgtcgtg aaggaagcag ttcctctgga agcttcttga agacaaacaa
     5281 cgtctgtagc gaccctttgc aggcagcgga accccccacc tggcgacagg tgcctctgcg
     5341 gccaaaagcc acgtgtataa gatacacctg caaaggcggc acaaccccag tgccacgttg
     5401 tgagttggat agttgtggaa agagtcaaat ggctctcctc aagcgtattc aacaaggggc
     5461 tgaaggatgc ccagaaggta ccccattgta tgggatctga tctggggcct cggtgcacat
     5521 gctttacatg tgtttagtcg aggttaaaaa aacgtctagg ccccccgaac cacggggacg
     5581 tggttttcct ttgaaaaaca cgatgataat atggccacaa ccatgaccga gtacaagccc
     5641 acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct cgccgccgcg
     5701 ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat cgagcgggtc
     5761 accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc
     5821 gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga agcgggggcg
     5881 gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct ggccgcgcag
     5941 caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc
     6001 accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc
     6061 ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacctc cgcgccccgc
     6121 aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa
     6181 ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gatgcatcga tgtcgacctg
     6241 cagccaagct tatcgataaa ataaaagatt ttatttagtc tccagaaaaa ggggggaatg
     6301 aaagacccca cctgtaggtt tggcaagcta gcttaagtaa cgccattttg caaggcatgg
     6361 aaaatacata actgagaata gagaagttca gatcaaggtt aggaacagag agacagcaga
     6421 atatgggcca aacaggatat ctgtggtaag cagttcctgc cccggctcag ggccaagaac
     6481 agatggtccc cagatgcggt cccgccctca gcagtttcta gagaaccatc agatgtttcc
     6541 agggtgcccc aaggacctga aatgaccctg tgccttattt gaactaacca atcagttcgc
     6601 ttctcgcttc tgttcgcgcg cttctgctcc ccgagctcaa taaaagagcc cacaacccct
     6661 cactcggcgc gccagtcctc cgatagactg cgtcgcccgg gtacccgtgt atccaataaa
     6721 ccctcttgca gttgcatccg acttgtggtc tcgctgttcc ttgggagggt ctcctctgag
     6781 tgattgacta cccgtcagcg ggggtctttc atgggtaaca gtttcttgaa gttggagaac
     6841 aacattctga gggtaggagt cgaatattaa gtaatcctga ctcaattagc cactgttttg
     6901 aatccacata ctccaatact cctgaaatag ttcattatgg acagcgcaga aagagctggg
     6961 gagaattgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     7021 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     7081 tgcccgcttt ccagtcggga aacctgtcgt gccagctg
//
