LOCUS       sequence-228323-        7526 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 7526)
  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 7526)
  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-USP46
FEATURES             Location/Qualifiers
     source          1..7526
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    579..920
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             987..1403
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1333..1355
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1371..1387
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      1416..1425
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1425..1448
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1461..1487
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    1503..1527
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2626..2650)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    2663..3230
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    2680..3232
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2824..2841)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3051..3070
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3244..3843
                     /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(3244..3263)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3740..3760
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             3919..4433
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    4595..4611
                     /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(4619..4625,4626..4643,4644..4649))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4664..4685
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4802..4819)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4973..5561)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5053..5072)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5732..6523,6524..6592))
                     /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     6355..6374
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6593..6697)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6765..6783
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6821..6843)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6943..6962
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7156..7178
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7306..7325
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(7525..7526,1..515)
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
ORIGIN
        1 tgaaagaccc cacctgtagg tttggcaagc tagcttaagt aacgccattt tgcaaggcat
       61 ggaaaataca taactgagaa tagagaagtt cagatcaagg ttaggaacag agagacagca
      121 gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc agggccaaga
      181 acagatggtc cccagatgcg gtcccgccct cagcagtttc tagagaacca tcagatgttt
      241 ccagggtgcc ccaaggacct gaaatgaccc tgtgccttat ttgaactaac caatcagttc
      301 gcttctcgct tctgttcgcg cgcttctgct ccccgagctc aataaaagag cccacaaccc
      361 ctcactcggc gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt attcccaata
      421 aagcctcttg ctgtttgcat ccgaatcgtg gactcgctga tccttgggag ggtctcctca
      481 gattgattga ctgcccacct cgggggtctt tcatttggag gttccaccga gatttggaga
      541 cccctgccca gggaccaccg acccccccgc cgggaggtaa gctggccagc ggtcgtttcg
      601 tgtctgtctc tgtctttgtg cgtgtttgtg ccggcatcta atgtttgcgc ctgcgtctgt
      661 actagttagc taactagctc tgtatctggc ggacccgtgg tggaactgac gagttctgaa
      721 cacccggccg caaccctggg agacgtccca gggactttgg gggccgtttt tgtggcccga
      781 cctgaggaag ggagtcgatg tggaatccga ccccgtcagg atatgtggtt ctggtaggag
      841 acgagaacct aaaacagttc ccgcctccgt ctgaattttt gctttcggtt tggaaccgaa
      901 gccgcgcgtc ttgtctgctg cagcgctgca gcatcgttct gtgttgtctc tgtctgactg
      961 tgtttctgta tttgtctgaa aattagggcc agactgttac cactccctta agtttgacct
     1021 taggtcactg gaaagatgtc gagcggatcg ctcacaacca gtcggtagat gtcaagaaga
     1081 gacgttgggt taccttctgc tctgcagaat ggccaacctt taacgtcgga tggccgcgag
     1141 acggcacctt taaccgagac ctcatcaccc aggttaagat caaggtcttt tcacctggcc
     1201 cgcatggaca cccagaccag gtcccctaca tcgtgacctg ggaagccttg gcttttgacc
     1261 cccctccctg ggtcaagccc tttgtacacc ctaagcctcc gcctcctctt cctccatccg
     1321 ccccgtctct cccccttgaa cctcctcgtt cgaccccgcc tcgatcctcc ctttatccag
     1381 ccctcactcc ttctctaggc gccggaatta gatctgccac catggactac aaggatgacg
     1441 atgacaagct cgatggagga tacccatacg atgttccaga ttacgctgct cgaggttatc
     1501 aaacaagttt gtacaaaaaa gttggcatga ctgtccgaaa catcgcctcc atctgtaata
     1561 tgggcaccaa tgcctctgct ctggaaaaag acattggtcc agagcagttt ccaatcaatg
     1621 aacactattt cggattggtc aattttggaa acacatgcta ctgtaactcc gtgcttcagg
     1681 cattgtactt ctgccgtcca ttccgggaga atgtgttggc atacaaggcc cagcaaaaga
     1741 agaaggaaaa cttgctgacg tgcctggcgg accttttcca cagcattgcc acacagaaga
     1801 agaaggttgg cgtcatccca ccaaagaagt tcatttcaag gctgagaaaa gagaatgatc
     1861 tctttgataa ctacatgcag caggatgctc atgaattttt aaattatttg ctaaacacta
     1921 ttgcggacat ccttcaggag gagaagaaac aggaaaaaca aaatggaaaa ttaaaaaatg
     1981 gcaacatgaa cgaacctgcg gaaaataata aaccagaact cacctgggtc catgagattt
     2041 ttcagggaac gcttaccaat gaaactcgat gcttgaactg tgaaactgtt agtagcaaag
     2101 atgaagattt tcttgacctt tctgttgatg tggagcagaa tacatccatt acccactgtc
     2161 taagagactt cagcaacaca gaaacactgt gtagtgaaca aaaatattat tgtgaaacat
     2221 gctgcagcaa acaagaagcc cagaaaagga tgagggtaaa aaagctgccc atgatcttgg
     2281 ccctgcacct aaagcggttc aagtacatgg agcagctgca cagatacacc aagctgtctt
     2341 accgtgtggt cttccctctg gaactccggc tcttcaacac ctccagtgat gcagtgaacc
     2401 tggaccgcat gtatgacttg gttgcggtgg tcgttcactg tggcagtggt cctaatcgtg
     2461 ggcattatat cactattgtg aaaagtcacg gcttctggct tttgtttgat gatgacattg
     2521 tagagaaaat agatgctcaa gctattgaag aattctatgg cctgacgtca gatatatcaa
     2581 aaaattcaga atctggatat attttattct atcagtcaag agagtaccca actttcttgt
     2641 acaaagtggt tcgataacga attccgcccc ccccccctaa cgttactggc cgaagccgct
     2701 tggaataagg ccggtgtgcg tttgtctata tgttattttc caccatattg ccgtcttttg
     2761 gcaatgtgag ggcccggaaa cctggccctg tcttcttgac gagcattcct aggggtcttt
     2821 cccctctcgc caaaggaatg caaggtctgt tgaatgtcgt gaaggaagca gttcctctgg
     2881 aagcttcttg aagacaaaca acgtctgtag cgaccctttg caggcagcgg aaccccccac
     2941 ctggcgacag gtgcctctgc ggccaaaagc cacgtgtata agatacacct gcaaaggcgg
     3001 cacaacccca gtgccacgtt gtgagttgga tagttgtgga aagagtcaaa tggctctcct
     3061 caagcgtatt caacaagggg ctgaaggatg cccagaaggt accccattgt atgggatctg
     3121 atctggggcc tcggtgcaca tgctttacat gtgtttagtc gaggttaaaa aaacgtctag
     3181 gccccccgaa ccacggggac gtggttttcc tttgaaaaac acgatgataa tatggccaca
     3241 accatgaccg agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt ccccagggcc
     3301 gtacgcaccc tcgccgccgc gttcgccgac taccccgcca cgcgccacac cgtcgatccg
     3361 gaccgccaca tcgagcgggt caccgagctg caagaactct tcctcacgcg cgtcgggctc
     3421 gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg
     3481 gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc
     3541 ggttcccggc tggccgcgca gcaacagatg gaaggcctcc tggcgccgca ccggcccaag
     3601 gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg accaccaggg caagggtctg
     3661 ggcagcgccg tcgtgctccc cggagtggag gcggccgagc gcgccggggt gcccgccttc
     3721 ctggagacct ccgcgccccg caacctcccc ttctacgagc ggctcggctt caccgtcacc
     3781 gccgacgtcg aggtgcccga aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc
     3841 tgatgcatcg atgtcgacct gcagccaagc ttatcgataa aataaaagat tttatttagt
     3901 ctccagaaaa aggggggaat gaaagacccc acctgtaggt ttggcaagct agcttaagta
     3961 acgccatttt gcaaggcatg gaaaatacat aactgagaat agagaagttc agatcaaggt
     4021 taggaacaga gagacagcag aatatgggcc aaacaggata tctgtggtaa gcagttcctg
     4081 ccccggctca gggccaagaa cagatggtcc ccagatgcgg tcccgccctc agcagtttct
     4141 agagaaccat cagatgtttc cagggtgccc caaggacctg aaatgaccct gtgccttatt
     4201 tgaactaacc aatcagttcg cttctcgctt ctgttcgcgc gcttctgctc cccgagctca
     4261 ataaaagagc ccacaacccc tcactcggcg cgccagtcct ccgatagact gcgtcgcccg
     4321 ggtacccgtg tatccaataa accctcttgc agttgcatcc gacttgtggt ctcgctgttc
     4381 cttgggaggg tctcctctga gtgattgact acccgtcagc gggggtcttt catgggtaac
     4441 agtttcttga agttggagaa caacattctg agggtaggag tcgaatatta agtaatcctg
     4501 actcaattag ccactgtttt gaatccacat actccaatac tcctgaaata gttcattatg
     4561 gacagcgcag aaagagctgg ggagaattgt gaaattgtta tccgctcaca attccacaca
     4621 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     4681 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     4741 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     4801 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     4861 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     4921 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     4981 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     5041 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     5101 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     5161 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     5221 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     5281 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     5341 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     5401 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     5461 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     5521 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     5581 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     5641 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     5701 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     5761 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     5821 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     5881 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     5941 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     6001 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     6061 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     6121 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     6181 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     6241 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     6301 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     6361 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     6421 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     6481 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     6541 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     6601 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     6661 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     6721 ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga
     6781 ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc
     6841 cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg
     6901 cgtcagcggg tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg
     6961 tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc
     7021 gcatcaggcg ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg
     7081 cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg
     7141 taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggcgcaagga atggtgcatg
     7201 caaggagatg gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa
     7261 acaagcgctc atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat
     7321 ataggcgcca gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta
     7381 gaggcgatta gtccaatttg ttaaagacag gatatcagtg gtccaggctc tagttttgac
     7441 tcaacaatat caccagctga agcctataga gtacgagcca tagataaaat aaaagatttt
     7501 atttagtctc cagaaaaagg ggggaa
//
