LOCUS       sequence-224850-        5934 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Production of recombinant Ufd1-His in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5934)
  AUTHORS   Weith M, Seiler J, van den Boom J, Kracht M, Hulsmann J, Primorac I,
            Del Pino Garcia J, Kaschani F, Kaiser M, Musacchio A, Bollen M, 
            Meyer H
  TITLE     Ubiquitin-Independent Disassembly by a p97 AAA-ATPase Complex Drives
            PP1 Holoenzyme Formation.
  JOURNAL   Mol Cell. 2018 Oct 11. pii: S1097-2765(18)30791-3. doi: 
            10.1016/j.molcel.2018.09.020.
  PUBMED    30344098
REFERENCE   2  (bases 1 to 5934)
  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. 
            2018 Oct 11. pii: S1097-2765(18)30791-3. doi: 
            10.1016/j.molcel.2018.09.020."
COMMENT     Sequence Label: pET41b+_Ufd1-His
FEATURES             Location/Qualifiers
     source          1..5934
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     70..89
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     177..196
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        177..195
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    196..220
                     /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             235..257
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1192..1215
                     /codon_start=1
                     /product="8xHis affinity tag"
                     /label=8xHis
                     /translation="HHHHHHHH"
     primer_bind     complement(1278..1296)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1292..1339
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1376..1831
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1463..1482)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1673..1694
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1923..2738)
                     /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(1923..2738)
                     /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     2646..2665
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2730..2749
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2860..3448
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3349..3368
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3602..3619
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3634..3773
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3699..3718)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3859..3881
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3875..4066)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4638..5720)
                     /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     5682..5701
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5721..5798)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 gcctgccacc atacccacgc cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc
       61 ttccccatcg gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg cgccggtgat
      121 gccggccacg atgcgtccgg cgtagaggat cgagatcgat ctcgatcccg cgaaattaat
      181 acgactcact ataggggaat tgtgagcgga taacaattcc cctctagaaa taattttgtt
      241 taactttaag aaggagatat acatatgttc tctttcaaca tgttcgacca ccctattccc
      301 agggtcttcc aaaaccgctt ctccacacag taccgctgct tctctgtgtc catgctagca
      361 gggcctaatg acaggtcaga tgtggagaaa ggagggaaga taattatgcc accctcggcc
      421 ctggaccaac tcagccgact taacattacc tatcccatgc tgttcaaact gaccaataag
      481 aattcggacc gcatgacgca ttgtggcgtg ctggagtttg tggctgatga gggcatctgc
      541 tacctcccac actggatgat gcagaactta ctcttggaag aaggcggcct ggtccaggtg
      601 gagagcgtca accttcaagt ggccacctac tccaaattcc aacctcagag ccctgacttc
      661 ctggacatca ccaaccccaa agccgtatta gaaaacgcac ttaggaactt tgcctgtctg
      721 accaccgggg atgtgattgc catcaactat aatgaaaaga tctacgaact gcgtgtgatg
      781 gagaccaaac ccgacaaggc agtgtccatc attgagtgtg acatgaacgt ggactttgat
      841 gctcccctgg gctacaaaga acccgaaaga caagtccagc atgaggagtc gacagaaggt
      901 gaagccgacc acagtggcta tgctggagag ctgggcttcc gcgctttctc tggatctggc
      961 aatagactgg atggaaagaa gaaaggggta gagcccagcc cctccccaat caagcctgga
     1021 gatattaaaa gaggaattcc caattatgaa tttaaacttg gtaagataac tttcatcaga
     1081 aattcacgtc cccttgtcaa aaaggttgaa gaggatgaag ctggaggcag attcgtcgct
     1141 ttctctggag aaggacagtc attgcgtaaa aagggaagaa agcccctcga gcaccaccac
     1201 caccaccacc accactaatt gattaatacc taggctgcta aacaaagccc gaaaggaagc
     1261 tgagttggct gctgccaccg ctgagcaata actagcataa ccccttgggg cctctaaacg
     1321 ggtcttgagg ggttttttgc tgaaaggagg aactatatcc ggattggcga atgggacgcg
     1381 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     1441 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     1501 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     1561 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     1621 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     1681 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     1741 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     1801 aattttaaca aaatattaac gcttacaatt taggtggcac ttttcgggga aatgtgcgcg
     1861 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgaattaat
     1921 tcttagaaaa actcatcgag catcaaatga aactgcaatt tattcatatc aggattatca
     1981 ataccatatt tttgaaaaag ccgtttctgt aatgaaggag aaaactcacc gaggcagttc
     2041 cataggatgg caagatcctg gtatcggtct gcgattccga ctcgtccaac atcaatacaa
     2101 cctattaatt tcccctcgtc aaaaataagg ttatcaagtg agaaatcacc atgagtgacg
     2161 actgaatccg gtgagaatgg caaaagttta tgcatttctt tccagacttg ttcaacaggc
     2221 cagccattac gctcgtcatc aaaatcactc gcatcaacca aaccgttatt cattcgtgat
     2281 tgcgcctgag cgagacgaaa tacgcgatcg ctgttaaaag gacaattaca aacaggaatc
     2341 gaatgcaacc ggcgcaggaa cactgccagc gcatcaacaa tattttcacc tgaatcagga
     2401 tattcttcta atacctggaa tgctgttttc ccggggatcg cagtggtgag taaccatgca
     2461 tcatcaggag tacggataaa atgcttgatg gtcggaagag gcataaattc cgtcagccag
     2521 tttagtctga ccatctcatc tgtaacatca ttggcaacgc tacctttgcc atgtttcaga
     2581 aacaactctg gcgcatcagg cttcccatac aatcgataga ttgtcgcacc tgattgcccg
     2641 acattatcgc gagcccattt atacccatat aaatcagcat ccatgttgga atttaatcgc
     2701 ggcctagagc aagacgtttc ccgttgaata tggctcataa caccccttgt attactgttt
     2761 atgtaagcag acagttttat tgttcatgac caaaatccct taacgtgagt tttcgttcca
     2821 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     2881 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     2941 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     3001 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     3061 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     3121 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     3181 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     3241 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     3301 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     3361 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     3421 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     3481 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     3541 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     3601 cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca
     3661 tctgtgcggt atttcacacc gcaatggtgc actctcagta caatctgctc tgatgccgca
     3721 tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac
     3781 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     3841 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     3901 aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat tcacagatgt
     3961 ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat gtctggcttc
     4021 tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat gcctccgtgt
     4081 aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg atgctcacga
     4141 tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt aaacaactgg
     4201 cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta
     4261 atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag atccggaaca
     4321 taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga
     4381 ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct
     4441 cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct agccgggtcc
     4501 tcaacgacag gagcacgatc atgctagtca tgccccgcgc ccaccggaag gagctgactg
     4561 ggttgaaggc tctcaagggc atcggtcgag atcccggtgc ctaatgagtg agctaactta
     4621 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     4681 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc cagggtggtt
     4741 tttcttttca ccagtgagac gggcaacagc tgattgccct tcaccgcctg gccctgagag
     4801 agttgcagca agcggtccac gctggtttgc cccagcaggc gaaaatcctg tttgatggtg
     4861 gttaacggcg ggatataaca tgagctgtct tcggtatcgt cgtatcccac taccgagatg
     4921 tccgcaccaa cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga
     4981 tcgttggcaa ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt
     5041 tgaaaaccgg acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg
     5101 cgagtgagat atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggt
     5161 cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc
     5221 gtaccgtctt catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga
     5281 aataacgccg gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc
     5341 ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta
     5401 caggcttcga cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg
     5461 gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg
     5521 gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg
     5581 taattcagct ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg
     5641 gcctggttca ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg
     5701 tataacgtta ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat
     5761 gccataccgc gaaaggtttt gcgccattcg atggtgtccg ggatctcgac gctctccctt
     5821 atgcgactcc tgcattagga agcagcccag tagtaggttg aggccgttga gcaccgccgc
     5881 cgcaaggaat ggtgcatgca aggagatggc gcccaacagt cccccggcca cggg
//
