LOCUS       sequence-350013-        6712 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Bacterial Expression of TPR2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6712)
  AUTHORS   Ke J, Ma H, Gu X, Thelen A, Brunzelle JS, Li J, Xu HE, Melcher K
  TITLE     Structural basis for recognition of diverse transcriptional 
            repressors by the TOPLESS family of corepressors.
  JOURNAL   Sci Adv. 2015 Jul 24;1(6):e1500107. doi: 10.1126/sciadv.1500107. 
            eCollection 2015 Jul.
  PUBMED    26601214
REFERENCE   2  (bases 1 to 6712)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Adv."; 
            date: "2015-07-24"; volume: "1(6):e1500107. doi"; pages: " 
            10.1126/sciadv.1500107. eCollection 2015 Jul"
COMMENT     Sequence Label: pSumo-His6-SUMO-OsTPR2(1-210)
FEATURES             Location/Qualifiers
     source          1..6712
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     969..988
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1008..1085)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1291..1310
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1394..1413
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1394..1412
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1413..1437
                     /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             1452..1474
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1487..1504
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1508..1798
                     /codon_start=1
                     /gene="S. cerevisiae SMT3 (truncated)"
                     /product="cleavable ubiquitin-like protein tag"
                     /label=SUMO
                     /translation="SDSEVNQEAKPEVKPEVKPETHINLKVSDGSSEIFFKIKKTTPLR
                     RLMEAFAKRQGKEMDSLRFLYDGIRIQADQAPEDLDMEDNDIIEAHREQIGG"
     CDS             2837..2854
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2907..2925)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2921..2968
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3005..3460
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3092..3111)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3302..3323
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3486..3590
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3591..3659,3660..4451)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3809..3828)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4622..5210
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5111..5130
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5364..5381
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5396..5535
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5461..5480)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5621..5643
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5637..5828)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6637..6712,1..1007))
                     /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 gtttgcgtat tgggcgccag ggtggttttt cttttcacca gtgagacggg caacagctga
       61 ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc
      121 agcaggcgaa aatcctgttt gatggtggtt aacggcggga tataacatga gctgtcttcg
      181 gtatcgtcgt atcccactac cgagatatcc gcaccaacgc gcagcccgga ctcggtaatg
      241 gcgcgcattg cgcccagcgc catctgatcg ttggcaacca gcatcgcagt gggaacgatg
      301 ccctcattca gcatttgcat ggtttgttga aaaccggaca tggcactcca gtcgccttcc
      361 cgttccgcta tcggctgaat ttgattgcga gtgagatatt tatgccagcc agccagacgc
      421 agacgcgccg agacagaact taatgggccc gctaacagcg cgatttgctg gtgacccaat
      481 gcgaccagat gctccacgcc cagtcgcgta ccgtcttcat gggagaaaat aatactgttg
      541 atgggtgtct ggtcagagac atcaagaaat aacgccggaa cattagtgca ggcagcttcc
      601 acagcaatgg catcctggtc atccagcgga tagttaatga tcagcccact gacgcgttgc
      661 gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc cgcttcgttc taccatcgac
      721 accaccacgc tggcacccag ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac
      781 ggcgcgtgca gggccagact ggaggtggca acgccaatca gcaacgactg tttgcccgcc
      841 agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg ccatcgccgc ttccactttt
      901 tcccgcgttt tcgcagaaac gtggctggcc tggttcacca cgcgggaaac ggtctgataa
      961 gagacaccgg catactctgc gacatcgtat aacgttactg gtttcacatt caccaccctg
     1021 aattgactct cttccgggcg ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg
     1081 gtgtccggga tctcgacgct ctcccttatg cgactcctgc attaggaagc agcccagtag
     1141 taggttgagg ccgttgagca ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc
     1201 caacagtccc ccggccacgg ggcctgccac catacccacg ccgaaacaag cgctcatgag
     1261 cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg gcgatatagg cgccagcaac
     1321 cgcacctgtg gcgccggtga tgccggccac gatgcgtccg gcgtagagga tcgagatctc
     1381 gatcccgcga aattaatacg actcactata ggggaattgt gagcggataa caattcccct
     1441 ctagaaataa ttttgtttaa ctttaagaag gagatatacc atgggtcatc accatcatca
     1501 tcacgggtcg gactcagaag tcaatcaaga agctaagcca gaggtcaagc cagaagtcaa
     1561 gcctgagact cacatcaatt taaaggtgtc cgatggatct tcagagatct tcttcaagat
     1621 caaaaagacc actcctttaa gaaggctgat ggaagcgttc gctaaaagac agggtaagga
     1681 aatggactcc ttaagattct tgtacgacgg tattagaatt caagctgatc aggcccctga
     1741 agatttggac atggaggata acgatattat tgaggctcac cgcgaacaga ttggaggtgg
     1801 atccatgtct tcattgtcaa gagagttggt cttcttaatt ttgcaattct tggacgagga
     1861 aaagttcaag gagacagtcc ataagttaga gcaggaatct ggattcttct tcaatatgaa
     1921 atacttcgag gagaaggtcc acgcaggtga gtgggacgag gttgagaagt acttgtcagg
     1981 atttactaag gttgatgata ataggtactc tatgaagata tttttcgaaa ttaggaaaca
     2041 aaagtattta gaggctttgg ataggcacga cagggctaag gctgtcgata tattggtcaa
     2101 ggacttgaaa gttttctcaa cttttaatga agaattgtat aaggaaataa ctcagttatt
     2161 gactttagag aatttcagag aaaatgagca gttgtctaag tacggtgata ctaagtctgc
     2221 tagatctatt atgttgatag aattgaaaaa gttaatagag gcaaacccat tgttcaggga
     2281 gaagttggtc ttccctactt taaaggcatc aaggttgagg acattgatta atcagtcatt
     2341 gaactggcag catcagttgt gcaagaatcc aaggccaaat cctgatataa agactttgtt
     2401 cactgaccac acatgtacac ctccaaacgg tgcaagggca tcaccagtct cagttccatt
     2461 ggcagcagtc ccaaaggcag gtggtacata ccctccattg acagcacaca ctcctttcca
     2521 gccacctcca gctggtcctt cattagcagg atggatggca aacgcagctg ctgcaacttc
     2581 atctgtccct tcagctgtcg ttgctgcatc ttcattgcct gtcccaccaa accaagcagt
     2641 ccctattatg aaaagaccaa caattactga ctaccaatct gctgaatcag aacagttgat
     2701 gaagaggtta agaccttcag gacacggtgt tgatgaggct acataccctg ctcctatacc
     2761 acagccattg tggtctgttg aggacttgcc aagaacagtt gcttgcactt tgtcacaggg
     2821 tgcggccgca ctcgagcacc accaccacca ccactgagat ccggctgcta acaaagcccg
     2881 aaaggaagct gagttggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc
     2941 ctctaaacgg gtcttgaggg gttttttgct gaaaggagga actatatccg gattggcgaa
     3001 tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     3061 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     3121 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     3181 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     3241 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     3301 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     3361 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     3421 tttaacgcga attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa
     3481 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     3541 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     3601 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     3661 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     3721 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     3781 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg
     3841 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     3901 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     3961 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     4021 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     4081 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgcagcaa
     4141 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     4201 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     4261 cggctggctg gtttattgct gataaatctg gagccggtga gcgtggttct cgcggtatca
     4321 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     4381 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     4441 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc
     4501 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     4561 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     4621 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     4681 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     4741 tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact
     4801 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     4861 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     4921 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     4981 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     5041 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     5101 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     5161 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     5221 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     5281 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
     5341 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga
     5401 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcaatggt gcactctcag
     5461 tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac
     5521 tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     5581 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     5641 aggttttcac cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg
     5701 tcgtgaagcg attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc
     5761 agaagcgtta atgtctggct tctgataaag cgggccatgt taagggcggt tttttcctgt
     5821 ttggtcactg atgcctccgt gtaaggggga tttctgttca tgggggtaat gataccgatg
     5881 aaacgagaga ggatgctcac gatacgggtt actgatgatg aacatgcccg gttactggaa
     5941 cgttgtgagg gtaaacaact ggcggtatgg atgcggcggg accagagaaa aatcactcag
     6001 ggtcaatgcc agcgcttcgt taatacagat gtaggtgttc cacagggtag ccagcagcat
     6061 cctgcgatgc agatccggaa cataatggtg cagggcgctg acttccgcgt ttccagactt
     6121 tacgaaacac ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga cgttttgcag
     6181 cagcagtcgc ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa
     6241 ccccgccagc ctagccgggt cctcaacgac aggagcacga tcatgcgcac ccgtggggcc
     6301 gccatgccgg cgataatggc ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg
     6361 aaggcttgag cgagggcgtg caagattccg aataccgcaa gcgacaggcc gatcatcgtc
     6421 gcgctccagc gaaagcggtc ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct
     6481 acgagttgca tgataaagaa gacagtcata agtgcggcga cgatagtcat gccccgcgcc
     6541 caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc
     6601 taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     6661 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cg
//
