LOCUS       sequence-157778-        6818 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6818)
  AUTHORS   Sheng Y, Laister RC, Lemak A, Wu B, Tai E, Duan S, Lukin J, 
            Sunnerhagen M, Srisailam S, Karra M, Benchimol S, Arrowsmith CH
  TITLE     Molecular basis of Pirh2-mediated p53 ubiquitylation.
  JOURNAL   Nat Struct Mol Biol. 2008 Dec . 15(12):1334-42.
  PUBMED    19043414
REFERENCE   2  (bases 1 to 6818)
  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: "Nat Struct 
            Mol Biol. 2008 Dec . 15(12):1334-42."
COMMENT     Sequence Label: p53-(1-393)-E285K-N268D
FEATURES             Location/Qualifiers
     source          1..6818
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..15,16..32,33..38)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     terminator      complement(213..260)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(264..1445)
                     /codon_start=1
                     /gene="Homo sapiens TP53"
                     /product="human tumor suppressor protein that is often 
                     mutated in cancers"
                     /label=p53
                     /note="responds to cellular stresses to regulate the 
                     expression of target genes"
                     /translation="MEEPQSDPSVEPPLSQETFSDLWKLLPENNVLSPLPSQAMDDLML
                     SPDDIEQWFTEDPGPDEAPRMPEAAPPVAPAPAAPTPAAPAPAPSWPLSSSVPSQKTYQ
                     GSYGFRLGFLHSGTAKSVTCTYSPALNKMFCQLAKTCPVQLWVDSTPPPGTRVRAMAIY
                     KQSQHMTEVVRRCPHHERCSDSDGLAPPQHLIRVEGNLRVEYLDDRNTFRHSVVVPYEP
                     PEVGSDCTTIHYNYMCNSSCMGGMNRRPILTIITLEDSSGNLLGRDSFEVRVCACPGRD
                     RRTKEENLRKKGEPHHELPPGSTKRALPNNTSSSPQPKKKPLDGEYFTLQIRGRERFEM
                     FRELNEALELKDAQAGKEPGGSRAHSSHLKSKKGQSTSRHKKLMFKTEGPDSD"
     CDS             complement(1449..1466)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(1476..1493)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     protein_bind    1546..1570
                     /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     complement(1570..1589)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1571..1589)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1673..1692)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1898..1975
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1976..3058
                     /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(1995..2014)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4370..4561
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(4555..4577)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4663..4805
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4718..4737
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4820..4837)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4991..5579)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5071..5090)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5750..6541,6542..6610))
                     /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     6373..6392
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6611..6715)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6783..6801
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ttctcatgtt tgacagctta tcatcgataa gctttaatgc ggtagtttat cacagttaaa
       61 ttgctaacgc agtcaggcac cgtgtatgaa atctaacaat gcgctcatcg tcatcctcgg
      121 caccgtcacc ctggatgctg taggcatagg cttggttatg ccggtactgc cgggcctctt
      181 gcgggatatc cggatatagt tcctcctttc agcaaaaaac ccctcaagac ccgtttagag
      241 gccccaaggg gttatgctag ttatcagtct gagtcaggcc cttctgtctt gaacatgagt
      301 tttttatggc gggaggtaga ctgacccttt ttggacttca ggtggctgga gtgagccctg
      361 ctcccccctg gctccttccc agcctgggca tccttgagtt ccaaggcctc attcagctct
      421 cggaacatct cgaagcgctc acgcccacgg atctgaaggg tgaaatattc tccatccagt
      481 ggtttcttct ttggctgggg agaggagctg gtgttgttgg gcagtgctcg cttagtgctc
      541 cctgggggca gctcgtggtg aggctcccct ttcttgcgga gattctcttc ctttgtgcgc
      601 cggtctctcc caggacaggc acaaacacgc acctcaaagc tgtcccgtcc cagtagatta
      661 ccactggagt cttccagtgt gatgatggtg aggatgggcc tccggttcat gccgcccatg
      721 caggaactgt tacacatgta gttgtagtgg atggtggtac agtcagagcc aacctcaggc
      781 ggctcatagg gcaccaccac actatgtcga aaagtgtttc tgtcatccaa atactccaca
      841 cgcaaatttc cttccactcg gataagatgc tgaggagggg ccagaccatc gctatctgag
      901 cagcgctcat ggtgggggca gcgcctcaca acctccgtca tgtgctgtga ctgcttgtag
      961 atggccatgg cgcggacgcg ggtgccgggc gggggtgtgg aatcaaccca cagctgcaca
     1021 gggcaggtct tggccagttg gcaaaacatc ttgttgaggg caggggagta cgtgcaagtc
     1081 acagacttgg ctgtcccaga atgcaagaag cccagacgga aaccgtagct gccctggtag
     1141 gttttctggg aagggacaga agatgacagg ggccaggagg gggctggtgc aggggccgcc
     1201 ggtgtaggag ctgctggtgc aggggccacg gggggagcag cctctggcat tctgggagct
     1261 tcatctggac ctgggtcttc ggtgaaccat tgttcaatat cgtccgggga cagcatcaaa
     1321 tcatccattg cttgggacgg caagggggac agaacgttgt tttcaggaag tagtttccat
     1381 aggtctgaaa atgtttcctg actcagaggg ggctcgacgc taggatctga ctgcggctcc
     1441 tccatatggc tgccgcgcgg caccaggccg ctgctgtgat gatgatgatg atggctgctg
     1501 cccatggtat atctccttct taaagtaaac aaaatattct agaggggaat tgttatccgc
     1561 tcacaattcc cctatagtga gtcgtattaa tttcgcggga tcgagatctc gatcctctac
     1621 gccggacgca tcgtggccgg catcaccggc gccacaggtg cggttgctgg cgcctatatc
     1681 gccgacatca ccgatgggga agatcgggct cgccacttcg ggctcatgag cgcttgtttc
     1741 ggcgtgggta tggtggcagg ccccgtggcc gggggactgt tgggcgccat ctccttgcat
     1801 gcaccattcc ttgcggcggc ggtgctcaac ggcctcaacc tactactggg ctgcttccta
     1861 atgcaggagt cgcataaggg agagcgtcga gatcccggac accatcgaat ggcgcaaaac
     1921 ctttcgcggt atggcatgat agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa
     1981 accagtaacg ttatacgatg tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg
     2041 cgtggtgaac caggccagcc acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat
     2101 ggcggagctg aattacattc ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt
     2161 gctgattggc gttgccacct ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc
     2221 gattaaatct cgcgccgatc aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag
     2281 cggcgtcgaa gcctgtaaag cggcggtgca caatcttctc gcgcaacgcg tcagtgggct
     2341 gatcattaac tatccgctgg atgaccagga tgccattgct gtggaagctg cctgcactaa
     2401 tgttccggcg ttatttcttg atgtctctga ccagacaccc atcaacagta ttattttctc
     2461 ccatgaagac ggtacgcgac tgggcgtgga gcatctggtc gcattgggtc accagcaaat
     2521 cgcgctgtta gcgggcccat taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca
     2581 taaatatctc actcgcaatc aaattcagcc gatagcggaa cgggaaggcg actggagtgc
     2641 catgtccggt tttcaacaaa ccatgcaaat gctgaatgag ggcatcgttc ccactgcgat
     2701 gctggttgcc aacgatcaga tggcgctggg cgcaatgcgc gccattaccg agtccgggct
     2761 gcgcgttggt gcggatatct cggtagtggg atacgacgat accgaagaca gctcatgtta
     2821 tatcccgccg ttaaccacca tcaaacagga ttttcgcctg ctggggcaaa ccagcgtgga
     2881 ccgcttgctg caactctctc agggccaggc ggtgaagggc aatcagctgt tgcccgtctc
     2941 actggtgaaa agaaaaacca ccctggcgcc caatacgcaa accgcctctc cccgcgcgtt
     3001 ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc
     3061 gcaacgcaat taatgtaagt tagctcactc attaggcacc gggatctcga ccgatgccct
     3121 tgagagcctt caacccagtc agctccttcc ggtgggcgcg gggcatgact atcgtcgccg
     3181 cacttatgac tgtcttcttt atcatgcaac tcgtaggaca ggtgccggca gcgctctggg
     3241 tcattttcgg cgaggaccgc tttcgctgga gcgcgacgat gatcggcctg tcgcttgcgg
     3301 tattcggaat cttgcacgcc ctcgctcaag ccttcgtcac tggtcccgcc accaaacgtt
     3361 tcggcgagaa gcaggccatt atcgccggca tggcggccga cgcgctgggc tacgtcttgc
     3421 tggcgttcgc gacgcgaggc tggatggcct tccccattat gattcttctc gcttccggcg
     3481 gcatcgggat gcccgcgttg caggccatgc tgtccaggca ggtagatgac gaccatcagg
     3541 gacagcttca aggatcgctc gcggctctta ccagcctaac ttcgatcact ggaccgctga
     3601 tcgtcacggc gatttatgcc gcctcggcga gcacatggaa cgggttggca tggattgtag
     3661 gcgccgccct ataccttgtc tgcctccccg cgttgcgtcg cggtgcatgg agccgggcca
     3721 cctcgacctg aatggaagcc ggcggcacct cgctaacgga ttcaccactc caagaattgg
     3781 agccaatcaa ttcttgcgga gaactgtgaa tgcgcaaacc aacccttggc agaacatatc
     3841 catcgcgtcc gccatctcca gcagccgcac gcggcgcatc tcgggcagcg ttgggtcctg
     3901 gccacgggtg cgcatgatcg tgctcctgtc gttgaggacc cggctaggct ggcggggttg
     3961 ccttactggt tagcagaatg aatcaccgat acgcgagcga acgtgaagcg actgctgctg
     4021 caaaacgtct gcgacctgag caacaacatg aatggtcttc ggtttccgtg tttcgtaaag
     4081 tctggaaacg cggaagtcag cgccctgcac cattatgttc cggatctgca tcgcaggatg
     4141 ctgctggcta ccctgtggaa cacctacatc tgtattaacg aagcgctggc attgaccctg
     4201 agtgattttt ctctggtccc gccgcatcca taccgccagt tgtttaccct cacaacgttc
     4261 cagtaaccgg gcatgttcat catcagtaac ccgtatcgtg agcatcctct ctcgtttcat
     4321 cggtatcatt acccccatga acagaaatcc cccttacacg gaggcatcag tgaccaaaca
     4381 ggaaaaaacc gcccttaaca tggcccgctt tatcagaagc cagacattaa cgcttctgga
     4441 gaaactcaac gagctggacg cggatgaaca ggcagacatc tgtgaatcgc ttcacgacca
     4501 cgctgatgag ctttaccgca gctgcctcgc gcgtttcggt gatgacggtg aaaacctctg
     4561 acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg ggagcagaca
     4621 agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggcgcagcca tgacccagtc
     4681 acgtagcgat agcggagtgt atactggctt aactatgcgg catcagagca gattgtactg
     4741 agagtgcacc atatatgcgg tgtgaaatac cgcacagatg cgtaaggaga aaataccgca
     4801 tcaggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     4861 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     4921 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     4981 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     5041 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     5101 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     5161 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     5221 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     5281 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     5341 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     5401 agtggtggcc taactacggc tacactagaa ggacagtatt tggtatctgc gctctgctga
     5461 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     5521 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     5581 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     5641 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     5701 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     5761 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     5821 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     5881 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     5941 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     6001 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     6061 ttgctgcagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     6121 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     6181 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     6241 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     6301 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     6361 cgtcaacacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     6421 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     6481 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     6541 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     6601 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     6661 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     6721 ttccccgaaa agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata
     6781 aaaataggcg tatcacgagg ccctttcgtc ttcaagaa
//
