LOCUS       sequence-456528-        6217 bp    DNA     circular SYN 14-APR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6217)
  AUTHORS   Wu CJ, Conze DB, Li T, Srinivasula SM, Ashwell JD
  TITLE     NEMO is a sensor of Lys 63-linked polyubiquitination and functions 
            in NF-kappaB activation.
  JOURNAL   Nat Cell Biol. 2006 Apr . 8(4):398-406.
  PUBMED    16547522
REFERENCE   2  (bases 1 to 6217)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat Cell 
            Biol. 2006 Apr . 8(4):398-406."
COMMENT     Sequence Label: pGEX-NEMO
FEATURES             Location/Qualifiers
     source          1..6217
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(166..171,172..187,188..194)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    202..218
                     /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             241..894
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     852..874
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             901..918
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(2250..2272)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2417..2435)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2503..2607
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2608..2676,2677..3468)
                     /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(2826..2845)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3639..4227
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4128..4147
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4381..4398
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        4471..4548
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4549..5631
                     /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(4568..4587)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(5680..5685,5686..5703,5704..5710)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5718..5734
                     /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     5742..5758
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5742..5758
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(5754..6217,1..64)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(5774..5791)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5774..5790)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5783..5805)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 gtgcaccaat gcttctggcg tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc
       61 gtaaatcact gcataattcg tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt
      121 gcgccgacat cataacggtt ctggcaaata ttctgaaatg agctgttgac aattaatcat
      181 cggctcgtat aatgtgtgga attgtgagcg gataacaatt tcacacagga aacagtattc
      241 atgtccccta tactaggtta ttggaaaatt aagggccttg tgcaacccac tcgacttctt
      301 ttggaatatc ttgaagaaaa atatgaagag catttgtatg agcgcgatga aggtgataaa
      361 tggcgaaaca aaaagtttga attgggtttg gagtttccca atcttcctta ttatattgat
      421 ggtgatgtta aattaacaca gtctatggcc atcatacgtt atatagctga caagcacaac
      481 atgttgggtg gttgtccaaa agagcgtgca gagatttcaa tgcttgaagg agcggttttg
      541 gatattagat acggtgtttc gagaattgca tatagtaaag actttgaaac tctcaaagtt
      601 gattttctta gcaagctacc tgaaatgctg aaaatgttcg aagatcgttt atgtcataaa
      661 acatatttaa atggtgatca tgtaacccat cctgacttca tgttgtatga cgctcttgat
      721 gttgttttat acatggaccc aatgtgcctg gatgcgttcc caaaattagt ttgttttaaa
      781 aaacgtattg aagctatccc acaaattgat aagtacttga aatccagcaa gtatatagca
      841 tggcctttgc agggctggca agccacgttt ggtggtggcg accatcctcc aaaatcggat
      901 ctggttccgc gtggatcccc ggaattcaat aggcacctct ggaagagcca actgtgtgag
      961 atggtgcagc ccagtggtgg cccggcagca gatcaggacg tactgggcga agagtctcct
     1021 ctggggaagc cagccatgct gcacctgcct tcagaacagg gcgctcctga gaccctccag
     1081 cgctgcctgg aggagaatca agagctccga gatgccatcc ggcagagcaa ccagattctg
     1141 cgggagcgct gcgaggagct tctgcatttc caagccagcc agagggagga gaaggagttc
     1201 ctcatgtgca agttccagga ggccaggaaa ctggtggaga gactcggcct ggagaagctc
     1261 gatctgaaga ggcagaagga gcaggctctg cgggaggtgg agcacctgaa gagatgccag
     1321 cagcagatgg ctgaggacaa ggcctctgtg aaagcccagg tgacgtcctt gctcggggag
     1381 ctgcaggaga gccagagtcg cttggaggct gccactaagg aatgccaggc tctggagggt
     1441 cgggcccggg cggccagcga gcaggcgcgg cagctggaga gtgagcgcga ggcgctgcag
     1501 cagcagcaca gcgtgcaggt ggaccagctg cgcatgcagg gccagagcgt ggaggccgcg
     1561 ctccgcatgg agcgccaggc cgcctcggag gagaagagga agctggccca gttgcaggtg
     1621 gcctatcacc agctcttcca agaatacgac aaccacatca agagcagcgt ggtgggcagt
     1681 gagcggaagc gaggaatgca gctggaagat ctcaaacagc agctccagca ggccgaggag
     1741 gccctggtgg ccaaacagga ggtgatcgat aagctgaagg aggaggccga gcagcacaag
     1801 attgtgatgg agaccgttcc ggtgctgaag gcccaggcgg atatctacaa ggcggacttc
     1861 caggctgaga ggcaggcccg ggagaagctg gccgagaaga aggagctcct gcaggagcag
     1921 ctggagcagc tgcagaggga gtacagcaaa ctgaaggcca gctgtcagga gtcggccagg
     1981 atcgaggaca tgaggaagcg gcatgtcgag gtctcccagg cccccttgcc ccccgcccct
     2041 gcctacctct cctctcccct ggccctgccc agccagagga ggagcccccc cgaggagcca
     2101 cctgacttct gctgtcccaa gtgccagtat caggcccctg atatggacac cctgcagata
     2161 catgtcatgg agtgcattga gtagctcgag cggccgcatc gtgactgact gacgatctgc
     2221 ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggagacggtc
     2281 acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt
     2341 gttggcgggt gtcggggcgc agccatgacc cagtcacgta gcgatagcgg agtgtataat
     2401 tcttgaagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata
     2461 atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     2521 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     2581 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     2641 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     2701 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     2761 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     2821 gttctgctat gtggcgcggt attatcccgt gttgacgccg ggcaagagca actcggtcgc
     2881 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     2941 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     3001 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     3061 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     3121 ccaaacgacg agcgtgacac cacgatgcct gcagcaatgg caacaacgtt gcgcaaacta
     3181 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     3241 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     3301 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     3361 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     3421 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     3481 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     3541 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     3601 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     3661 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     3721 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     3781 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     3841 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     3901 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     3961 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     4021 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     4081 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     4141 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     4201 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     4261 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     4321 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     4381 agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat
     4441 ctgtgcggta tttcacaccg cataaattcc gacaccatcg aatggcgcaa aacctttcgc
     4501 ggtatggcat gatagcgccc ggaagagagt caattcaggg tggtgaatgt gaaaccagta
     4561 acgttatacg atgtcgcaga gtatgccggt gtctcttatc agaccgtttc ccgcgtggtg
     4621 aaccaggcca gccacgtttc tgcgaaaacg cgggaaaaag tggaagcggc gatggcggag
     4681 ctgaattaca ttcccaaccg cgtggcacaa caactggcgg gcaaacagtc gttgctgatt
     4741 ggcgttgcca cctccagtct ggccctgcac gcgccgtcgc aaattgtcgc ggcgattaaa
     4801 tctcgcgccg atcaactggg tgccagcgtg gtggtgtcga tggtagaacg aagcggcgtc
     4861 gaagcctgta aagcggcggt gcacaatctt ctcgcgcaac gcgtcagtgg gctgatcatt
     4921 aactatccgc tggatgacca ggatgccatt gctgtggaag ctgcctgcac taatgttccg
     4981 gcgttatttc ttgatgtctc tgaccagaca cccatcaaca gtattatttt ctcccatgaa
     5041 gacggtacgc gactgggcgt ggagcatctg gtcgcattgg gtcaccagca aatcgcgctg
     5101 ttagcgggcc cattaagttc tgtctcggcg cgtctgcgtc tggctggctg gcataaatat
     5161 ctcactcgca atcaaattca gccgatagcg gaacgggaag gcgactggag tgccatgtcc
     5221 ggttttcaac aaaccatgca aatgctgaat gagggcatcg ttcccactgc gatgctggtt
     5281 gccaacgatc agatggcgct gggcgcaatg cgcgccatta ccgagtccgg gctgcgcgtt
     5341 ggtgcggata tctcggtagt gggatacgac gataccgaag acagctcatg ttatatcccg
     5401 ccgttaacca ccatcaaaca ggattttcgc ctgctggggc aaaccagcgt ggaccgcttg
     5461 ctgcaactct ctcagggcca ggcggtgaag ggcaatcagc tgttgcccgt ctcactggtg
     5521 aaaagaaaaa ccaccctggc gcccaatacg caaaccgcct ctccccgcgc gttggccgat
     5581 tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc
     5641 aattaatgta agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc
     5701 tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca
     5761 tgattacgga ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta
     5821 cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg
     5881 cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg cgctttgcct
     5941 ggtttccggc accagaagcg gtgccggaaa gctggctgga gtgcgatctt cctgaggccg
     6001 atactgtcgt cgtcccctca aactggcaga tgcacggtta cgatgcgccc atctacacca
     6061 acgtgaccta tcccattacg gtcaatccgc cgtttgttcc cacggagaat ccgacgggtt
     6121 gttactcgct cacatttaat gttgatgaaa gctggctaca ggaaggccag acgcgaatta
     6181 tttttgatgg cgttggaatt agcttatcga ctgcacg
//
