LOCUS       sequence-259537-        6074 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  expresses human RNF146 full length in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6074)
  AUTHORS   DaRosa PA, Wang Z, Jiang X, Pruneda JN, Cong F, Klevit RE, Xu W
  TITLE     Allosteric activation of the RNF146 ubiquitin ligase by a 
            poly(ADP-ribosyl)ation signal.
  JOURNAL   Nature. 2015 Jan 8;517(7533):223-6. doi: 10.1038/nature13826. Epub 
            2014 Oct 19.
  PUBMED    25327252
REFERENCE   2  (bases 1 to 6074)
  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"; doi: 
            "10.1038/nature13826"; journalName: "Nature"; date: "2015-01-8- 8"; 
            volume: "517"; issue: "7533"; pages: "223-6"
COMMENT     Sequence Label: pGEX6P-2-human RNF146 full length
FEATURES             Location/Qualifiers
     source          1..6074
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..17
                     /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)."
     CDS             40..693
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     651..673
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             700..723
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             1819..1836
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1906..1928)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2073..2091)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2159..2263
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2264..2332,2333..3124)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2482..2501)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3295..3883
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3784..3803
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4037..4054
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4127..4204
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4205..5287
                     /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     complement(4224..4243)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5336..5341,5342..5359,5360..5366)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5374..5390
                     /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)."
     primer_bind     5398..5414
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5398..5414
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             5410..5937
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5430..5447)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5430..5446)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5439..5461)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        join(6039..6044,6045..6060,6061..6067)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
ORIGIN
        1 ttgtgagcgg ataacaattt cacacaggaa acagtattca tgtcccctat actaggttat
       61 tggaaaatta agggccttgt gcaacccact cgacttcttt tggaatatct tgaagaaaaa
      121 tatgaagagc atttgtatga gcgcgatgaa ggtgataaat ggcgaaacaa aaagtttgaa
      181 ttgggtttgg agtttcccaa tcttccttat tatattgatg gtgatgttaa attaacacag
      241 tctatggcca tcatacgtta tatagctgac aagcacaaca tgttgggtgg ttgtccaaaa
      301 gagcgtgcag agatttcaat gcttgaagga gcggttttgg atattagata cggtgtttcg
      361 agaattgcat atagtaaaga ctttgaaact ctcaaagttg attttcttag caagctacct
      421 gaaatgctga aaatgttcga agatcgttta tgtcataaaa catatttaaa tggtgatcat
      481 gtaacccatc ctgacttcat gttgtatgac gctcttgatg ttgttttata catggaccca
      541 atgtgcctgg atgcgttccc aaaattagtt tgttttaaaa aacgtattga agctatccca
      601 caaattgata agtacttgaa atccagcaag tatatagcat ggcctttgca gggctggcaa
      661 gccacgtttg gtggtggcga ccatcctcca aaatcggatc tggaagttct gttccagggg
      721 cccctgggat ccccaggaat tcatatggct ggctgtggtg aaattgatca ttcaataaac
      781 atgcttccta caaacaggaa agcgaacgag tcctgttcta atactgcacc ttctttaacc
      841 gtccctgaat gtgccatttg tctgcaaaca tgtgttcatc cagtcagtct gccctgtaag
      901 cacgttttct gctatctatg tgtaaaagga gcttcatggc ttggaaagcg gtgtgctctt
      961 tgtcgacaag aaattcccga ggatttcctt gacaagccaa ccttgttgtc accagaagaa
     1021 ctcaaggcag caagtagagg aaatggtgaa tatgcatggt attatgaagg aagaaatggg
     1081 tggtggcagt acgatgagcg cactagtaga gagctggaag atgctttttc caaaggtaaa
     1141 aagaacactg aaatgttaat tgctggcttt ctgtatgtcg ctgatcttga aaacatggtt
     1201 caatatagga gaaatgaaca tggacgtcgc aggaagatta agcgagatat aatagatata
     1261 ccaaagaagg gagtagctgg acttaggcta gactgtgatg ctaataccgt aaacctagca
     1321 agagagagct ctgctgacgg agcggacagt gtatcagcac agagtggagc ttctgttcag
     1381 cccctagtgt cttctgtaag gcccctaaca tcagtagatg gtcagttaac aagccctgca
     1441 acaccatccc ctgatgcaag cacttctctg gaagactctt ttgctcattt acaactcagt
     1501 ggagacaaca cagctgaaag gagtcatagg ggagaaggag aagaagatca tgaatcacca
     1561 tcttcaggca gggtaccagc accagacacc tccattgaag aaactgaatc agatgccagt
     1621 agtgatagtg aggatgtatc tgcagttgtt gcacagcact ccttgaccca acagagactt
     1681 ttggtttcta atgcaaacca gacagtaccc gatcgatcag atcgatcggg aactgatcga
     1741 tcagtagcag ggggtggaac agtgagtgtc agtgtcagat ctagaaggcc tgatggacag
     1801 tgcacagtaa ctgaagttca ccaccaccac caccactaag ctcgagcggc cgcatcgtga
     1861 ctgactgacg atctgcctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     1921 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
     1981 gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga
     2041 tagcggagtg tataattctt gaagacgaaa gggcctcgtg atacgcctat ttttataggt
     2101 taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg
     2161 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     2221 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     2281 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     2341 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     2401 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     2461 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtgttg acgccgggca
     2521 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     2581 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     2641 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     2701 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     2761 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgcag caatggcaac
     2821 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     2881 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     2941 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     3001 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     3061 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     3121 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     3181 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     3241 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     3301 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     3361 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     3421 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     3481 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     3541 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     3601 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     3661 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     3721 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     3781 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     3841 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     3901 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     3961 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     4021 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta
     4081 ttttctcctt acgcatctgt gcggtatttc acaccgcata aattccgaca ccatcgaatg
     4141 gcgcaaaacc tttcgcggta tggcatgata gcgcccggaa gagagtcaat tcagggtggt
     4201 gaatgtgaaa ccagtaacgt tatacgatgt cgcagagtat gccggtgtct cttatcagac
     4261 cgtttcccgc gtggtgaacc aggccagcca cgtttctgcg aaaacgcggg aaaaagtgga
     4321 agcggcgatg gcggagctga attacattcc caaccgcgtg gcacaacaac tggcgggcaa
     4381 acagtcgttg ctgattggcg ttgccacctc cagtctggcc ctgcacgcgc cgtcgcaaat
     4441 tgtcgcggcg attaaatctc gcgccgatca actgggtgcc agcgtggtgg tgtcgatggt
     4501 agaacgaagc ggcgtcgaag cctgtaaagc ggcggtgcac aatcttctcg cgcaacgcgt
     4561 cagtgggctg atcattaact atccgctgga tgaccaggat gccattgctg tggaagctgc
     4621 ctgcactaat gttccggcgt tatttcttga tgtctctgac cagacaccca tcaacagtat
     4681 tattttctcc catgaagacg gtacgcgact gggcgtggag catctggtcg cattgggtca
     4741 ccagcaaatc gcgctgttag cgggcccatt aagttctgtc tcggcgcgtc tgcgtctggc
     4801 tggctggcat aaatatctca ctcgcaatca aattcagccg atagcggaac gggaaggcga
     4861 ctggagtgcc atgtccggtt ttcaacaaac catgcaaatg ctgaatgagg gcatcgttcc
     4921 cactgcgatg ctggttgcca acgatcagat ggcgctgggc gcaatgcgcg ccattaccga
     4981 gtccgggctg cgcgttggtg cggatatctc ggtagtggga tacgacgata ccgaagacag
     5041 ctcatgttat atcccgccgt taaccaccat caaacaggat tttcgcctgc tggggcaaac
     5101 cagcgtggac cgcttgctgc aactctctca gggccaggcg gtgaagggca atcagctgtt
     5161 gcccgtctca ctggtgaaaa gaaaaaccac gctagcgccc aatacgcaaa ccgcctctcc
     5221 ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg
     5281 gcagtgagcg caacgcaatt aatgtaagtt agctcactca ttaggcaccc caggctttac
     5341 actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa tttcacacag
     5401 gaaacagcta tgaccatgat tacggattca ctggccgtcg ttttacaacg tcgtgactgg
     5461 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg
     5521 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     5581 gaatggcgct ttgcctggtt tccggcacca gaagcggtgc cggaaagctg gctggagtgc
     5641 gatcttcctg aggccgatac tgtcgtcgtc ccctcaaact ggcagatgca cggttacgat
     5701 gcgcccatct acaccaacgt gacctatccc attacggtca atccgccgtt tgttcccacg
     5761 gagaatccga cgggttgtta ctcgctcaca tttaatgttg atgaaagctg gctacaggaa
     5821 ggccagacgc gaattatttt tgatggcgtt ggaattagct tatcgactgc acggtgcacc
     5881 aatgcttctg gcgtcaggca gccatcggaa gctgtggtat ggctgtgcag gtcgtaaatc
     5941 actgcataat tcgtgtcgct caaggcgcac tcccgttctg gataatgttt tttgcgccga
     6001 catcataacg gttctggcaa atattctgaa atgagctgtt gacaattaat catcggctcg
     6061 tataatgtgt ggaa
//
