LOCUS       sequence-394854-        5881 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  bacterial expression of P53 TET-CTD for segmental labeling.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5881)
  AUTHORS   Krois AS, Park S, Martinez-Yamout MA, Dyson HJ, Wright PE
  TITLE     Mapping Interactions of the Intrinsically Disordered C-Terminal 
            Regions of Tetrameric p53 by Segmental Isotope Labeling and NMR.
  JOURNAL   Biochemistry. 2022 Dec 6;61(23):2709-2719. doi: 
            10.1021/acs.biochem.2c00528. Epub 2022 Nov 15.
  PUBMED    36380579
REFERENCE   2  (bases 1 to 5881)
  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: 
            "10.1021/acs.biochem.2c00528"; journalName: "Biochemistry"; date: 
            "2022-12-6- 6"; volume: "61"; issue: "23"; pages: "2709-2719"
COMMENT     Sequence Label: H6SUMO-IntC-P53(304-393)
FEATURES             Location/Qualifiers
     source          1..5881
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     919..938
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(958..1035)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1241..1260
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1344..1363
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1344..1362
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1363..1387
                     /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             1402..1424
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1443..1460
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1470..1487
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             2089..2106
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      2173..2220
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2257..2712
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2344..2363)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2554..2575
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2804..3619)
                     /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(2804..3619)
                     /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     3527..3546
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3611..3630
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3741..4329
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4230..4249
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4483..4500
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4515..4654
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4580..4599)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4740..4762
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4756..4947)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(5756..5881,1..957))
                     /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 caacagctga ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct
       61 ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt aacggcggga tataacatga
      121 gctgtcttcg gtatcgtcgt atcccactac cgagatatcc gcaccaacgc gcagcccgga
      181 ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg ttggcaacca gcatcgcagt
      241 gggaacgatg ccctcattca gcatttgcat ggtttgttga aaaccggaca tggcactcca
      301 gtcgccttcc cgttccgcta tcggctgaat ttgattgcga gtgagatatt tatgccagcc
      361 agccagacgc agacgcgccg agacagaact taatgggccc gctaacagcg cgatttgctg
      421 gtgacccaat gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat
      481 aatactgttg atgggtgtct ggtcagagac atcaagaaat aacgccggaa cattagtgca
      541 ggcagcttcc acagcaatgg catcctggtc atccagcgga tagttaatga tcagcccact
      601 gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc
      661 taccatcgac accaccacgc tggcacccag ttgatcggcg cgagatttaa tcgccgcgac
      721 aatttgcgac ggcgcgtgca gggccagact ggaggtggca acgccaatca gcaacgactg
      781 tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc
      841 ttccactttt tcccgcgttt tcgcagaaac gtggctggcc tggttcacca cgcgggaaac
      901 ggtctgataa gagacaccgg catactctgc gacatcgtat aacgttactg gtttcacatt
      961 caccaccctg aattgactct cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg
     1021 ccattcgatg gtgtccggga tctcgacgct ctcccttatg cgactcctgc attaggaagc
     1081 agcccagtag taggttgagg ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg
     1141 agatggcgcc caacagtccc ccggccacgg ggcctgccac catacccacg ccgaaacaag
     1201 cgctcatgag cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg gcgatatagg
     1261 cgccagcaac cgcacctgtg gcgccggtga tgccggccac gatgcgtccg gcgtagagga
     1321 tcgagatctc gatcccgcga aattaatacg actcactata ggggaattgt gagcggataa
     1381 caattcccct ctagaaataa ttttgtttaa ctttaagaag gagatatacc atgggcagca
     1441 gccatcatca tcatcatcac agcagcggcc tggtgccgcg cggcagcgag gtcaagccag
     1501 aagtcaagcc tgagactcac atcaatttaa aggtgtccga tggatcttca gagatcttct
     1561 tcaagatcaa aaagaccact cctttaagaa ggctgatgga agcgttcgct aaaagacagg
     1621 gtaaggaaat ggactcctta agattcttgt acgacggtat tagaattcaa gctgatcaga
     1681 cccctgaaga tttggacatg gaggataacg atattattga ggctcacaga gaacagattg
     1741 gtggtgacca taattttgca ctcaaaaatg gcttcatagc ttctaattgt ttcaatggta
     1801 ccaagcgagc actgcccaac aacaccagct cctctcccca gccaaagaag aaaccactgg
     1861 atggagaata tttcaccctt cagatccgtg ggcgtgagcg cttcgagatg ttccgagagc
     1921 tgaatgaggc cttggaactc aaggatgccc aggctgggaa ggagccaggg gggagcaggg
     1981 ctcactccag ccacctgaag tccaaaaagg gtcagtctac ctcccgccat aaaaaactca
     2041 tgttcaagac agaagggcct gactcagact aatgcggccg cactcgagca ccaccaccac
     2101 caccactgag atccggctgc taacaaagcc cgaaaggaag ctgagttggc tgctgccgct
     2161 gctgagcaat aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg
     2221 ctgaaaggag gaactatatc cggattggcg aatgggacgc gccctgtagc ggcgcattaa
     2281 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     2341 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     2401 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     2461 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     2521 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     2581 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     2641 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     2701 cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     2761 tttctaaata cattcaaata tgtatccgct catgaattaa ttcttagaaa aactcatcga
     2821 gcatcaaatg aaactgcaat ttattcatat caggattatc aataccatat ttttgaaaaa
     2881 gccgtttctg taatgaagga gaaaactcac cgaggcagtt ccataggatg gcaagatcct
     2941 ggtatcggtc tgcgattccg actcgtccaa catcaataca acctattaat ttcccctcgt
     3001 caaaaataag gttatcaagt gagaaatcac catgagtgac gactgaatcc ggtgagaatg
     3061 gcaaaagttt atgcatttct ttccagactt gttcaacagg ccagccatta cgctcgtcat
     3121 caaaatcact cgcatcaacc aaaccgttat tcattcgtga ttgcgcctga gcgagacgaa
     3181 atacgcgatc gctgttaaaa ggacaattac aaacaggaat cgaatgcaac cggcgcagga
     3241 acactgccag cgcatcaaca atattttcac ctgaatcagg atattcttct aatacctgga
     3301 atgctgtttt cccggggatc gcagtggtga gtaaccatgc atcatcagga gtacggataa
     3361 aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca gtttagtctg accatctcat
     3421 ctgtaacatc attggcaacg ctacctttgc catgtttcag aaacaactct ggcgcatcgg
     3481 gcttcccata caatcgatag attgtcgcac ctgattgccc gacattatcg cgagcccatt
     3541 tatacccata taaatcagca tccatgttgg aatttaatcg cggcctagag caagacgttt
     3601 cccgttgaat atggctcata acaccccttg tattactgtt tatgtaagca gacagtttta
     3661 ttgttcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     3721 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     3781 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     3841 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag
     3901 ccatagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     3961 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     4021 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     4081 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     4141 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     4201 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     4261 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     4321 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     4381 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     4441 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     4501 gaagcggaag agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     4561 cgcaatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac
     4621 tccgctatcg ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga
     4681 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     4741 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg
     4801 gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc
     4861 cagctcgttg agtttctcca gaagcgttaa tgtctggctt ctgataaagc gggccatgtt
     4921 aagggcggtt ttttcctgtt tggtcactga tgcctccgtg taagggggat ttctgttcat
     4981 gggggtaatg ataccgatga aacgagagag gatgctcacg atacgggtta ctgatgatga
     5041 acatgcccgg ttactggaac gttgtgaggg taaacaactg gcggtatgga tgcggcggga
     5101 ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc
     5161 acagggtagc cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgctga
     5221 cttccgcgtt tccagacttt acgaaacacg gaaaccgaag accattcatg ttgttgctca
     5281 ggtcgcagac gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt
     5341 ctgctaacca gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca ggagcacgat
     5401 catgcgcacc cgtggggccg ccatgccggc gataatggcc tgcttctcgc cgaaacgttt
     5461 ggtggcggga ccagtgacga aggcttgagc gagggcgtgc aagattccga ataccgcaag
     5521 cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag
     5581 cgctgccggc acctgtccta cgagttgcat gataaagaag acagtcataa gtgcggcgac
     5641 gatagtcatg ccccgcgccc accggaagga gctgactggg ttgaaggctc tcaagggcat
     5701 cggtcgagat cccggtgcct aatgagtgag ctaacttaca ttaattgcgt tgcgctcact
     5761 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     5821 ggggagaggc ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg
     5881 g
//
