LOCUS       sequence-237564-        7241 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 7241)
  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 7241)
  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-OTUB1
FEATURES             Location/Qualifiers
     source          1..7241
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             154..670
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
     misc_feature    734..1075
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             1142..1558
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1488..1510
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1526..1542
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      1571..1580
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1580..1603
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1616..1642
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    1658..1682
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2496..2520)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    2533..3100
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    2550..3102
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2694..2711)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2921..2940
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3114..3713
                     /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(3114..3133)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3610..3630
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             3789..4303
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    4465..4481
                     /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(4489..4495,4496..4513,4514..4519))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4534..4555
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4672..4689)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4843..5431)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4923..4942)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5602..6393,6394..6462))
                     /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     6225..6244
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6463..6567)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6635..6653
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6691..6713)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6813..6832
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7026..7048
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7176..7195
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 tccggcgtag aggcgattag tccaatttgt taaagacagg atatcagtgg tccaggctct
       61 agttttgact caacaatatc accagctgaa gcctatagag tacgagccat agataaaata
      121 aaagatttta tttagtctcc agaaaaaggg gggaatgaaa gaccccacct gtaggtttgg
      181 caagctagct taagtaacgc cattttgcaa ggcatggaaa atacataact gagaatagag
      241 aagttcagat caaggttagg aacagagaga cagcagaata tgggccaaac aggatatctg
      301 tggtaagcag ttcctgcccc ggctcagggc caagaacaga tggtccccag atgcggtccc
      361 gccctcagca gtttctagag aaccatcaga tgtttccagg gtgccccaag gacctgaaat
      421 gaccctgtgc cttatttgaa ctaaccaatc agttcgcttc tcgcttctgt tcgcgcgctt
      481 ctgctccccg agctcaataa aagagcccac aacccctcac tcggcgcgcc agtcctccga
      541 tagactgcgt cgcccgggta cccgtattcc caataaagcc tcttgctgtt tgcatccgaa
      601 tcgtggactc gctgatcctt gggagggtct cctcagattg attgactgcc cacctcgggg
      661 gtctttcatt tggaggttcc accgagattt ggagacccct gcccagggac caccgacccc
      721 cccgccggga ggtaagctgg ccagcggtcg tttcgtgtct gtctctgtct ttgtgcgtgt
      781 ttgtgccggc atctaatgtt tgcgcctgcg tctgtactag ttagctaact agctctgtat
      841 ctggcggacc cgtggtggaa ctgacgagtt ctgaacaccc ggccgcaacc ctgggagacg
      901 tcccagggac tttgggggcc gtttttgtgg cccgacctga ggaagggagt cgatgtggaa
      961 tccgaccccg tcaggatatg tggttctggt aggagacgag aacctaaaac agttcccgcc
     1021 tccgtctgaa tttttgcttt cggtttggaa ccgaagccgc gcgtcttgtc tgctgcagcg
     1081 ctgcagcatc gttctgtgtt gtctctgtct gactgtgttt ctgtatttgt ctgaaaatta
     1141 gggccagact gttaccactc ccttaagttt gaccttaggt cactggaaag atgtcgagcg
     1201 gatcgctcac aaccagtcgg tagatgtcaa gaagagacgt tgggttacct tctgctctgc
     1261 agaatggcca acctttaacg tcggatggcc gcgagacggc acctttaacc gagacctcat
     1321 cacccaggtt aagatcaagg tcttttcacc tggcccgcat ggacacccag accaggtccc
     1381 ctacatcgtg acctgggaag ccttggcttt tgacccccct ccctgggtca agccctttgt
     1441 acaccctaag cctccgcctc ctcttcctcc atccgccccg tctctccccc ttgaacctcc
     1501 tcgttcgacc ccgcctcgat cctcccttta tccagccctc actccttctc taggcgccgg
     1561 aattagatct gccaccatgg actacaagga tgacgatgac aagctcgatg gaggataccc
     1621 atacgatgtt ccagattacg ctgctcgagg ttatcaaaca agtttgtaca aaaaagttgg
     1681 catggcggcg gaggaacctc agcagcagaa gcaggagccg ctgggcagcg actccgaagg
     1741 tgttaactgt ctggcctatg atgaagccat catggctcag caggaccgaa ttcagcaaga
     1801 gattgctgtg cagaaccctc tggtgtcaga gcggctggag ctctcggtcc tatacaagga
     1861 gtatgctgaa gatgacaaca tctatcaaca gaagatcaag gacctccaca aaaagtactc
     1921 gtacatccgc aagaccaggc ctgacggcaa ctgtttctat cgggctttcg gattctccca
     1981 cttggaggca ctgctggatg acagcaagga gttgcagcgg ttcaaggctg tgtctgccaa
     2041 gagcaaggaa gacctggtgt cccagggctt cactgaattc acaattgagg atttccacaa
     2101 cacgttcatg gacctgattg agcaggtgga gaagcagacc tctgtcgccg acctgctggc
     2161 ctccttcaat gaccagagca cctccgacta ccttgtggtc tacctgcggc tgctcacctc
     2221 gggctacctg cagcgcgaga gcaagttctt cgagcacttc atcgagggtg gacggactgt
     2281 caaggagttc tgccagcagg aggtggagcc catgtgcaag gagagcgacc acatccacat
     2341 cattgcgctg gcccaggccc tcagcgtgtc catccaggtg gagtacatgg accgcggcga
     2401 gggcggcacc accaatccgc acatcttccc tgagggctcc gagcccaagg tctaccttct
     2461 ctaccggcct ggacactacg atatcctcta caaataccca actttcttgt acaaagtggt
     2521 tcgataacga attccgcccc ccccccctaa cgttactggc cgaagccgct tggaataagg
     2581 ccggtgtgcg tttgtctata tgttattttc caccatattg ccgtcttttg gcaatgtgag
     2641 ggcccggaaa cctggccctg tcttcttgac gagcattcct aggggtcttt cccctctcgc
     2701 caaaggaatg caaggtctgt tgaatgtcgt gaaggaagca gttcctctgg aagcttcttg
     2761 aagacaaaca acgtctgtag cgaccctttg caggcagcgg aaccccccac ctggcgacag
     2821 gtgcctctgc ggccaaaagc cacgtgtata agatacacct gcaaaggcgg cacaacccca
     2881 gtgccacgtt gtgagttgga tagttgtgga aagagtcaaa tggctctcct caagcgtatt
     2941 caacaagggg ctgaaggatg cccagaaggt accccattgt atgggatctg atctggggcc
     3001 tcggtgcaca tgctttacat gtgtttagtc gaggttaaaa aaacgtctag gccccccgaa
     3061 ccacggggac gtggttttcc tttgaaaaac acgatgataa tatggccaca accatgaccg
     3121 agtacaagcc cacggtgcgc ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc
     3181 tcgccgccgc gttcgccgac taccccgcca cgcgccacac cgtcgatccg gaccgccaca
     3241 tcgagcgggt caccgagctg caagaactct tcctcacgcg cgtcgggctc gacatcggca
     3301 aggtgtgggt cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg
     3361 aagcgggggc ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc ggttcccggc
     3421 tggccgcgca gcaacagatg gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt
     3481 ggttcctggc caccgtcggc gtctcgcccg accaccaggg caagggtctg ggcagcgccg
     3541 tcgtgctccc cggagtggag gcggccgagc gcgccggggt gcccgccttc ctggagacct
     3601 ccgcgccccg caacctcccc ttctacgagc ggctcggctt caccgtcacc gccgacgtcg
     3661 aggtgcccga aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc tgatgcatcg
     3721 atgtcgacct gcagccaagc ttatcgataa aataaaagat tttatttagt ctccagaaaa
     3781 aggggggaat gaaagacccc acctgtaggt ttggcaagct agcttaagta acgccatttt
     3841 gcaaggcatg gaaaatacat aactgagaat agagaagttc agatcaaggt taggaacaga
     3901 gagacagcag aatatgggcc aaacaggata tctgtggtaa gcagttcctg ccccggctca
     3961 gggccaagaa cagatggtcc ccagatgcgg tcccgccctc agcagtttct agagaaccat
     4021 cagatgtttc cagggtgccc caaggacctg aaatgaccct gtgccttatt tgaactaacc
     4081 aatcagttcg cttctcgctt ctgttcgcgc gcttctgctc cccgagctca ataaaagagc
     4141 ccacaacccc tcactcggcg cgccagtcct ccgatagact gcgtcgcccg ggtacccgtg
     4201 tatccaataa accctcttgc agttgcatcc gacttgtggt ctcgctgttc cttgggaggg
     4261 tctcctctga gtgattgact acccgtcagc gggggtcttt catgggtaac agtttcttga
     4321 agttggagaa caacattctg agggtaggag tcgaatatta agtaatcctg actcaattag
     4381 ccactgtttt gaatccacat actccaatac tcctgaaata gttcattatg gacagcgcag
     4441 aaagagctgg ggagaattgt gaaattgtta tccgctcaca attccacaca acatacgagc
     4501 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     4561 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     4621 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     4681 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     4741 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     4801 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     4861 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     4921 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     4981 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     5041 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     5101 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     5161 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     5221 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     5281 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     5341 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     5401 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     5461 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     5521 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     5581 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     5641 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     5701 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     5761 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     5821 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     5881 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     5941 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     6001 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     6061 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     6121 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     6181 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     6241 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     6301 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     6361 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     6421 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     6481 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     6541 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcta
     6601 agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg
     6661 tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc cggagacggt
     6721 cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg cgtcagcggg
     6781 tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg tactgagagt
     6841 gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg agaaaatacc gcatcaggcg
     6901 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
     6961 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
     7021 gttttcccag tcacgacgtt gtaaaacgac ggcgcaagga atggtgcatg caaggagatg
     7081 gcgcccaaca gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc
     7141 atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca
     7201 gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc g
//
