LOCUS       sequence-239456-        7483 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 7483)
  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 7483)
  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-EIF3S3
FEATURES             Location/Qualifiers
     source          1..7483
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     180..202
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     330..349
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             549..1065
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
     misc_feature    1129..1470
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             1537..1953
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1883..1905
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1921..1937
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      1966..1975
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1975..1998
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2011..2037
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    2053..2077
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3133..3157)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    3170..3737
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    3187..3739
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3331..3348)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3558..3577
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3751..4350
                     /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(3751..3770)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4247..4267
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4426..4940
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5102..5118
                     /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(5126..5132,5133..5150,5151..5156))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5171..5192
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5309..5326)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5480..6068)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5560..5579)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6239..7030,7031..7099))
                     /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     6862..6881
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7100..7204)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7272..7290
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7328..7350)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7450..7469
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tatgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgccattc
       61 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
      121 ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc
      181 ccagtcacga cgttgtaaaa cgacggcgca aggaatggtg catgcaagga gatggcgccc
      241 aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc gctcatgagc
      301 ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc gccagcaacc
      361 gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggcg attagtccaa
      421 tttgttaaag acaggatatc agtggtccag gctctagttt tgactcaaca atatcaccag
      481 ctgaagccta tagagtacga gccatagata aaataaaaga ttttatttag tctccagaaa
      541 aaggggggaa tgaaagaccc cacctgtagg tttggcaagc tagcttaagt aacgccattt
      601 tgcaaggcat ggaaaataca taactgagaa tagagaagtt cagatcaagg ttaggaacag
      661 agagacagca gaatatgggc caaacaggat atctgtggta agcagttcct gccccggctc
      721 agggccaaga acagatggtc cccagatgcg gtcccgccct cagcagtttc tagagaacca
      781 tcagatgttt ccagggtgcc ccaaggacct gaaatgaccc tgtgccttat ttgaactaac
      841 caatcagttc gcttctcgct tctgttcgcg cgcttctgct ccccgagctc aataaaagag
      901 cccacaaccc ctcactcggc gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt
      961 attcccaata aagcctcttg ctgtttgcat ccgaatcgtg gactcgctga tccttgggag
     1021 ggtctcctca gattgattga ctgcccacct cgggggtctt tcatttggag gttccaccga
     1081 gatttggaga cccctgccca gggaccaccg acccccccgc cgggaggtaa gctggccagc
     1141 ggtcgtttcg tgtctgtctc tgtctttgtg cgtgtttgtg ccggcatcta atgtttgcgc
     1201 ctgcgtctgt actagttagc taactagctc tgtatctggc ggacccgtgg tggaactgac
     1261 gagttctgaa cacccggccg caaccctggg agacgtccca gggactttgg gggccgtttt
     1321 tgtggcccga cctgaggaag ggagtcgatg tggaatccga ccccgtcagg atatgtggtt
     1381 ctggtaggag acgagaacct aaaacagttc ccgcctccgt ctgaattttt gctttcggtt
     1441 tggaaccgaa gccgcgcgtc ttgtctgctg cagcgctgca gcatcgttct gtgttgtctc
     1501 tgtctgactg tgtttctgta tttgtctgaa aattagggcc agactgttac cactccctta
     1561 agtttgacct taggtcactg gaaagatgtc gagcggatcg ctcacaacca gtcggtagat
     1621 gtcaagaaga gacgttgggt taccttctgc tctgcagaat ggccaacctt taacgtcgga
     1681 tggccgcgag acggcacctt taaccgagac ctcatcaccc aggttaagat caaggtcttt
     1741 tcacctggcc cgcatggaca cccagaccag gtcccctaca tcgtgacctg ggaagccttg
     1801 gcttttgacc cccctccctg ggtcaagccc tttgtacacc ctaagcctcc gcctcctctt
     1861 cctccatccg ccccgtctct cccccttgaa cctcctcgtt cgaccccgcc tcgatcctcc
     1921 ctttatccag ccctcactcc ttctctaggc gccggaatta gatctgccac catggactac
     1981 aaggatgacg atgacaagct cgatggagga tacccatacg atgttccaga ttacgctgct
     2041 cgaggttatc aaacaagttt gtacaaaaaa gttggcatgg catcccgcaa ggaaggtacc
     2101 ggctctactg ccacctcttc cagctccacc gccggcgcag cagggaaagg caaaggcaaa
     2161 ggcggctcgg gagattcagc cgtgaagcaa gtgcagatag atggccttgt ggtattaaag
     2221 ataatcaaac attatcaaga agaaggacaa ggaactgaag ttgttcaagg agtgcttttg
     2281 ggtctggttg tagaagatcg gcttgaaatt accaactgct ttcctttccc tcagcacaca
     2341 gaggatgatg ctgactttga tgaagtccaa tatcagatgg aaatgatgcg gagccttcgc
     2401 catgtaaaca ttgatcatct tcacgtgggc tggtatcagt ccacatacta tggctcattc
     2461 gttacccggg cactcctgga ctctcagttt agttaccagc atgccattga agaatctgtc
     2521 gttctcattt atgatcccat aaaaactgcc caaggatctc tctcactaaa ggcatacaga
     2581 ctgactccta aactgatgga agtttgtaaa gaaaaggatt tttcccctga agcattgaaa
     2641 aaagcaaata tcacctttga gtacatgttt gaagaagtgc cgattgtaat taaaaattca
     2701 catctgatca atgtcctaat gtgggaactt gaaaagaagt cagctgttgc agataaacat
     2761 gaattgctca gccttgccag cagcaatcat ttggggaaga atctacagtt gctgatggac
     2821 agagtggatg aaatgagcca agatatagtt aaatacaaca catacatgag gaatactagt
     2881 aaacaacagc agcagaaaca tcagtatcag cagcgtcgcc agcaggagaa tatgcagcgc
     2941 cagagccgag gagaaccccc gctccctgag gaggacctgt ccaaactctt caaaccacca
     3001 cagccgcctg ccaggatgga ctcgctgctc attgcaggcc agataaacac ttactgccag
     3061 aacatcaagg agttcactgc ccaaaactta ggcaagctct tcatggccca ggctcttcaa
     3121 gatacaacaa ctacccaact ttcttgtaca aagtggttcg ataacgaatt ccgccccccc
     3181 cccctaacgt tactggccga agccgcttgg aataaggccg gtgtgcgttt gtctatatgt
     3241 tattttccac catattgccg tcttttggca atgtgagggc ccggaaacct ggccctgtct
     3301 tcttgacgag cattcctagg ggtctttccc ctctcgccaa aggaatgcaa ggtctgttga
     3361 atgtcgtgaa ggaagcagtt cctctggaag cttcttgaag acaaacaacg tctgtagcga
     3421 ccctttgcag gcagcggaac cccccacctg gcgacaggtg cctctgcggc caaaagccac
     3481 gtgtataaga tacacctgca aaggcggcac aaccccagtg ccacgttgtg agttggatag
     3541 ttgtggaaag agtcaaatgg ctctcctcaa gcgtattcaa caaggggctg aaggatgccc
     3601 agaaggtacc ccattgtatg ggatctgatc tggggcctcg gtgcacatgc tttacatgtg
     3661 tttagtcgag gttaaaaaaa cgtctaggcc ccccgaacca cggggacgtg gttttccttt
     3721 gaaaaacacg atgataatat ggccacaacc atgaccgagt acaagcccac ggtgcgcctc
     3781 gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac
     3841 cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa
     3901 gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc
     3961 gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag
     4021 atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa
     4081 ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc
     4141 tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg
     4201 gccgagcgcg ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc
     4261 tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc
     4321 tggtgcatga cccgcaagcc cggtgcctga tgcatcgatg tcgacctgca gccaagctta
     4381 tcgataaaat aaaagatttt atttagtctc cagaaaaagg ggggaatgaa agaccccacc
     4441 tgtaggtttg gcaagctagc ttaagtaacg ccattttgca aggcatggaa aatacataac
     4501 tgagaataga gaagttcaga tcaaggttag gaacagagag acagcagaat atgggccaaa
     4561 caggatatct gtggtaagca gttcctgccc cggctcaggg ccaagaacag atggtcccca
     4621 gatgcggtcc cgccctcagc agtttctaga gaaccatcag atgtttccag ggtgccccaa
     4681 ggacctgaaa tgaccctgtg ccttatttga actaaccaat cagttcgctt ctcgcttctg
     4741 ttcgcgcgct tctgctcccc gagctcaata aaagagccca caacccctca ctcggcgcgc
     4801 cagtcctccg atagactgcg tcgcccgggt acccgtgtat ccaataaacc ctcttgcagt
     4861 tgcatccgac ttgtggtctc gctgttcctt gggagggtct cctctgagtg attgactacc
     4921 cgtcagcggg ggtctttcat gggtaacagt ttcttgaagt tggagaacaa cattctgagg
     4981 gtaggagtcg aatattaagt aatcctgact caattagcca ctgttttgaa tccacatact
     5041 ccaatactcc tgaaatagtt cattatggac agcgcagaaa gagctgggga gaattgtgaa
     5101 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     5161 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     5221 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     5281 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     5341 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     5401 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     5461 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     5521 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     5581 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     5641 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     5701 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     5761 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     5821 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     5881 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     5941 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     6001 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     6061 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     6121 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     6181 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     6241 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     6301 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     6361 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     6421 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     6481 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     6541 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     6601 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     6661 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     6721 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     6781 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     6841 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     6901 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     6961 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     7021 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     7081 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     7141 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     7201 cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga aaccattatt atcatgacat
     7261 taacctataa aaataggcgt atcacgaggc cctttcgtct cgcgcgtttc ggtgatgacg
     7321 gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg
     7381 ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggctggc
     7441 ttaactatgc ggcatcagag cagattgtac tgagagtgca cca
//
