LOCUS       sequence-471113-        6384 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Protein overexpression of human uroporphyrinogen decarboxylase 
            N367A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6384)
  TITLE     A method for the detection and enrichment of endogenous cereblon 
            substrates
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6384)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 7, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pET28a-His6-UROD-N367A
FEATURES             Location/Qualifiers
     source          1..6384
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     523..542
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(562..639)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     845..864
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     948..967
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        948..966
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    967..991
                     /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             1006..1028
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1039..1056
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1081..1101
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     primer_bind     complement(2266..2284)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2280..2327
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2364..2819
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2451..2470)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2661..2682
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2911..3726)
                     /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(2911..3726)
                     /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     3634..3653
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3718..3737
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3848..4436
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4337..4356
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4590..4607
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4622..4761
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4687..4706)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4847..4869
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4863..5054)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5863..6384,1..561))
                     /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"
ORIGIN
        1 gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg
       61 cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag
      121 aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag
      181 cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt
      241 acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc
      301 ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt
      361 ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat
      421 gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct
      481 ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc
      541 gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca
      601 tgccataccg cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct
      661 tatgcgactc ctgcattagg aagcagccca gtagtaggtt gaggccgttg agcaccgccg
      721 ccgcaaggaa tggtgcatgc aaggagatgg cgcccaacag tcccccggcc acggggcctg
      781 ccaccatacc cacgccgaaa caagcgctca tgagcccgaa gtggcgagcc cgatcttccc
      841 catcggtgat gtcggcgata taggcgccag caaccgcacc tgtggcgccg gtgatgccgg
      901 ccacgatgcg tccggcgtag aggatcgaga tctcgatccc gcgaaattaa tacgactcac
      961 tataggggaa ttgtgagcgg ataacaattc ccctctagaa ataattttgt ttaactttaa
     1021 gaaggagata tacatatgca tcaccatcac caccattcgt ccggagtcga cttaggcacc
     1081 gagaatttgt attttcagag catggaagcg aacggccttg gccctcaagg gtttccagag
     1141 ttgaaaaacg ataccttcct tcgtgcagcc tggggggaag aaacggacta tacacccgtg
     1201 tggtgtatgc gccaggctgg ccgttacctt ccagagttcc gcgaaacgcg tgcggcgcaa
     1261 gattttttta gcacctgccg ttctcccgag gcgtgctgcg aattgactct tcagccgtta
     1321 cgccgttttc cacttgatgc ggcgatcatc ttcagcgaca ttttggtagt ccctcaagct
     1381 ctgggtatgg aggtaacgat ggttccaggg aaggggccgt cctttccaga accactgcgc
     1441 gaagagcagg acttggaacg cttgcgtgac ccggaggtgg tggcgtccga gctggggtat
     1501 gtcttccagg ctattaccct tacacgccag cgtcttgcgg gacgtgttcc tctgattgga
     1561 tttgctggag caccatggac tcttatgacc tacatggtag agggaggagg cagctctact
     1621 atggcccagg caaaacgctg gttgtaccaa cgtccacaag cgtcgcacca attgttacgc
     1681 attctgaccg atgccttggt gccttatctt gtgggtcagg tcgtcgcagg tgcgcaggcc
     1741 ttgcagttat ttgagtcaca cgcaggtcac cttggaccac agctgtttaa caaatttgct
     1801 ctgccctata tccgtgatgt ggccaagcaa gttaaggccc gtttgcgcga ggcgggtctt
     1861 gcaccagtac ctatgatcat ctttgccaag gatgggcact tcgccttgga ggaattggca
     1921 caggcaggct acgaagtagt tgggcttgac tggacggtag ccccgaagaa agctcgtgag
     1981 tgcgtgggga agacggttac attgcagggg aatctggacc cctgtgcttt atatgctagc
     2041 gaagaggaaa tcggtcaact tgtcaaacaa atgttggacg actttggtcc acatcgctat
     2101 atcgcaaatc ttggccatgg actgtacccg gatatggatc cagagcacgt aggcgcattc
     2161 gttgatgccg tccataagca ctcccgttta ctgcgccaag cttaaaggtg gatacggatc
     2221 cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct gagcaataac
     2281 tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa
     2341 ctatatccgg attggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     2401 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     2461 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     2521 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     2581 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     2641 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     2701 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     2761 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta
     2821 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     2881 tcaaatatgt atccgctcat gaattaattc ttagaaaaac tcatcgagca tcaaatgaaa
     2941 ctgcaattta ttcatatcag gattatcaat accatatttt tgaaaaagcc gtttctgtaa
     3001 tgaaggagaa aactcaccga ggcagttcca taggatggca agatcctggt atcggtctgc
     3061 gattccgact cgtccaacat caatacaacc tattaatttc ccctcgtcaa aaataaggtt
     3121 atcaagtgag aaatcaccat gagtgacgac tgaatccggt gagaatggca aaagtttatg
     3181 catttctttc cagacttgtt caacaggcca gccattacgc tcgtcatcaa aatcactcgc
     3241 atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg agacgaaata cgcgatcgct
     3301 gttaaaagga caattacaaa caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc
     3361 atcaacaata ttttcacctg aatcaggata ttcttctaat acctggaatg ctgttttccc
     3421 ggggatcgca gtggtgagta accatgcatc atcaggagta cggataaaat gcttgatggt
     3481 cggaagaggc ataaattccg tcagccagtt tagtctgacc atctcatctg taacatcatt
     3541 ggcaacgcta cctttgccat gtttcagaaa caactctggc gcatcgggct tcccatacaa
     3601 tcgatagatt gtcgcacctg attgcccgac attatcgcga gcccatttat acccatataa
     3661 atcagcatcc atgttggaat ttaatcgcgg cctagagcaa gacgtttccc gttgaatatg
     3721 gctcataaca ccccttgtat tactgtttat gtaagcagac agttttattg ttcatgacca
     3781 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     3841 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     3901 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     3961 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     4021 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     4081 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     4141 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     4201 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     4261 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     4321 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     4381 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     4441 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     4501 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     4561 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     4621 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc aatggtgcac
     4681 tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc gctatcgcta
     4741 cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc gccctgacgg
     4801 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg
     4861 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta aagctcatca
     4921 gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag ctcgttgagt
     4981 ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag ggcggttttt
     5041 tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg ggtaatgata
     5101 ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca tgcccggtta
     5161 ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca gagaaaaatc
     5221 actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca gggtagccag
     5281 cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt ccgcgtttcc
     5341 agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt cgcagacgtt
     5401 ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg ctaaccagta
     5461 aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat gcgcacccgt
     5521 ggggccgcca tgccggcgat aatggcctgc ttctcgccga aacgtttggt ggcgggacca
     5581 gtgacgaagg cttgagcgag ggcgtgcaag attccgaata ccgcaagcga caggccgatc
     5641 atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga cccagagcgc tgccggcacc
     5701 tgtcctacga gttgcatgat aaagaagaca gtcataagtg cggcgacgat agtcatgccc
     5761 cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg tcgagatccc
     5821 ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc cgctttccag
     5881 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     5941 ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca acagctgatt
     6001 gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg tttgccccag
     6061 caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc tgtcttcggt
     6121 atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact cggtaatggc
     6181 gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc
     6241 ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt cgccttcccg
     6301 ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag ccagacgcag
     6361 acgcgccgag acagaactta atgg
//
