LOCUS       sequence-345354-        5896 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  UIMLx2 ubiquitin probe for K48 ubiquitylated proteins.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5896)
  AUTHORS   Villamil M, Xiao W, Yu C, Huang L, Xu P, Kaiser P
  TITLE     The Ubiquitin Interacting Motif-Like Domain of Met4 Selectively 
            Binds K48 Polyubiquitin Chains.
  JOURNAL   Mol Cell Proteomics. 2022 Jan;21(1):100175. doi: 
            10.1016/j.mcpro.2021.100175. Epub 2021 Nov 9.
  PUBMED    34763062
REFERENCE   2  (bases 1 to 5896)
  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: "Mol Cell 
            Proteomics."; date: "2022-01"; volume: "21(1):100175. doi"; pages: "
            10.1016/j.mcpro.2021.100175. Epub 2021 Nov 9"
COMMENT     Sequence Label: UIMLx2
FEATURES             Location/Qualifiers
     source          1..5896
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(39..1121)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     1083..1102
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1122..1199)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1405..1424
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1508..1527
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1508..1526
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1527..1551
                     /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)."
     RBS             1566..1588
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1601..1645
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             1652..1669
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1679..1696
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1700..1993
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="MSDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPL
                     RRLMEAFAKRQGKEMDSLRFLYDGISIQADQTPEDLDMEDNDIIEAHREQIGG"
     CDS             2268..2285
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2338..2356)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2352..2399
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2436..2891
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2523..2542)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2733..2754
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2983..3798)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2983..3798)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3706..3725
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3790..3809
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3920..4508
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4409..4428
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4662..4679
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4694..4833
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4759..4778)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4919..4941
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4935..5126)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 cctaatgagt gagctaactt acattaattg cgttgcgctc actgcccgct ttccagtcgg
       61 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
      121 gtattgggcg ccagggtggt ttttcttttc accagtgaga cgggcaacag ctgattgccc
      181 ttcaccgcct ggccctgaga gagttgcagc aagcggtcca cgctggtttg ccccagcagg
      241 cgaaaatcct gtttgatggt ggttaacggc gggatataac atgagctgtc ttcggtatcg
      301 tcgtatccca ctaccgagat atccgcacca acgcgcagcc cggactcggt aatggcgcgc
      361 attgcgccca gcgccatctg atcgttggca accagcatcg cagtgggaac gatgccctca
      421 ttcagcattt gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc
      481 gctatcggct gaatttgatt gcgagtgaga tatttatgcc agccagccag acgcagacgc
      541 gccgagacag aacttaatgg gcccgctaac agcgcgattt gctggtgacc caatgcgacc
      601 agatgctcca cgcccagtcg cgtaccgtct tcatgggaga aaataatact gttgatgggt
      661 gtctggtcag agacatcaag aaataacgcc ggaacattag tgcaggcagc ttccacagca
      721 atggcatcct ggtcatccag cggatagtta atgatcagcc cactgacgcg ttgcgcgaga
      781 agattgtgca ccgccgcttt acaggcttcg acgccgcttc gttctaccat cgacaccacc
      841 acgctggcac ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg
      901 tgcagggcca gactggaggt ggcaacgcca atcagcaacg actgtttgcc cgccagttgt
      961 tgtgccacgc ggttgggaat gtaattcagc tccgccatcg ccgcttccac tttttcccgc
     1021 gttttcgcag aaacgtggct ggcctggttc accacgcggg aaacggtctg ataagagaca
     1081 ccggcatact ctgcgacatc gtataacgtt actggtttca cattcaccac cctgaattga
     1141 ctctcttccg ggcgctatca tgccataccg cgaaaggttt tgcgccattc gatggtgtcc
     1201 gggatctcga cgctctccct tatgcgactc ctgcattagg aagcagccca gtagtaggtt
     1261 gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg cgcccaacag
     1321 tcccccggcc acggggcctg ccaccatacc cacgccgaaa caagcgctca tgagcccgaa
     1381 gtggcgagcc cgatcttccc catcggtgat gtcggcgata taggcgccag caaccgcacc
     1441 tgtggcgccg gtgatgccgg ccacgatgcg tctggcgtag aggatcgaga tctcgatccc
     1501 gcgaaattaa tacgactcac tataggggaa ttgtgagcgg ataacaattc ccctctagaa
     1561 ataattttgt ttaactttaa gaaggagata taccatgggc ggtctgaacg acatcttcga
     1621 ggctcagaaa atcgaatggc acgaaagcag ccatcatcat catcatcaca gcagcggcct
     1681 ggtgccgcgc ggcagccata tgtcggactc agaagtcaat caagaagcta agccagaggt
     1741 caagccagaa gtcaagcctg agactcacat caatttaaag gtgtccgatg gatcttcaga
     1801 gatcttcttc aagatcaaaa agaccactcc tttaagaagg ctgatggaag cgttcgctaa
     1861 aagacagggt aaggaaatgg actccttaag attcttgtac gacggtatta gcattcaagc
     1921 tgatcagacc cctgaagatt tggacatgga ggataacgat attattgagg ctcacagaga
     1981 acagattggt ggatccgaat tcgagctccg tcgacaaggc aacactaacc tatatgacca
     2041 cgctgtcacg ccagaaatac tcttagaaca gttggcctac gtggataact tcataccatc
     2101 tctggataac gagttctcta atgtggattg gggcggcggc ggcaacacta acctatatga
     2161 ccacgctgtc acgccagaaa tactcttaga acagttggcc tacgtggata acttcatacc
     2221 atctctggat aacgagttct ctaatgtgga ttggtgaggc tctcgagcac caccaccacc
     2281 accactgaga tccggctgct aacaaagccc gaaaggaagc tgagttggct gctgccaccg
     2341 ctgagcaata actagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgc
     2401 tgaaaggagg aactatatcc ggattggcga atgggacgcg ccctgtagcg gcgcattaag
     2461 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     2521 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     2581 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     2641 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     2701 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     2761 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     2821 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac
     2881 gcttacaatt taggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     2941 ttctaaatac attcaaatat gtatccgctc atgaattaat tcttagaaaa actcatcgag
     3001 catcaaatga aactgcaatt tattcatatc aggattatca ataccatatt tttgaaaaag
     3061 ccgtttctgt aatgaaggag aaaactcacc gaggcagttc cataggatgg caagatcctg
     3121 gtatcggtct gcgattccga ctcgtccaac atcaatacaa cctattaatt tcccctcgtc
     3181 aaaaataagg ttatcaagtg agaaatcacc atgagtgacg actgaatccg gtgagaatgg
     3241 caaaagttta tgcatttctt tccagacttg ttcaacaggc cagccattac gctcgtcatc
     3301 aaaatcactc gcatcaacca aaccgttatt cattcgtgat tgcgcctgag cgagacgaaa
     3361 tacgcgatcg ctgttaaaag gacaattaca aacaggaatc gaatgcaacc ggcgcaggaa
     3421 cactgccagc gcatcaacaa tattttcacc tgaatcagga tattcttcta atacctggaa
     3481 tgctgttttc ccggggatcg cagtggtgag taaccatgca tcatcaggag tacggataaa
     3541 atgcttgatg gtcggaagag gcataaattc cgtcagccag tttagtctga ccatctcatc
     3601 tgtaacatca ttggcaacgc tacctttgcc atgtttcaga aacaactctg gcgcatcggg
     3661 cttcccatac aatcgataga ttgtcgcacc tgattgcccg acattatcgc gagcccattt
     3721 atacccatat aaatcagcat ccatgttgga atttaatcgc ggcctagagc aagacgtttc
     3781 ccgttgaata tggctcataa caccccttgt attactgttt atgtaagcag acagttttat
     3841 tgttcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     3901 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     3961 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     4021 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
     4081 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     4141 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     4201 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     4261 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     4321 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     4381 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     4441 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     4501 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     4561 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     4621 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     4681 aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     4741 gcaatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agtatacact
     4801 ccgctatcgc tacgtgactg ggtcatggct gcgccccgac acccgccaac acccgctgac
     4861 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
     4921 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg
     4981 taaagctcat cagcgtggtc gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc
     5041 agctcgttga gtttctccag aagcgttaat gtctggcttc tgataaagcg ggccatgtta
     5101 agggcggttt tttcctgttt ggtcactgat gcctccgtgt aagggggatt tctgttcatg
     5161 ggggtaatga taccgatgaa acgagagagg atgctcacga tacgggttac tgatgatgaa
     5221 catgcccggt tactggaacg ttgtgagggt aaacaactgg cggtatggat gcggcgggac
     5281 cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta atacagatgt aggtgttcca
     5341 cagggtagcc agcagcatcc tgcgatgcag atccggaaca taatggtgca gggcgctgac
     5401 ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag
     5461 gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc
     5521 tgctaaccag taaggcaacc ccgccagcct agccgggtcc tcaacgacag gagcacgatc
     5581 atgcgcaccc gtggggccgc catgccggcg ataatggcct gcttctcgcc gaaacgtttg
     5641 gtggcgggac cagtgacgaa ggcttgagcg agggcgtgca agattccgaa taccgcaagc
     5701 gacaggccga tcatcgtcgc gctccagcga aagcggtcct cgccgaaaat gacccagagc
     5761 gctgccggca cctgtcctac gagttgcatg ataaagaaga cagtcataag tgcggcgacg
     5821 atagtcatgc cccgcgccca ccggaaggag ctgactgggt tgaaggctct caagggcatc
     5881 ggtcgagatc ccggtg
//
