LOCUS       pET28a-ThrRS            7175 bp    DNA     circular SYN 22-JUL-2025
DEFINITION  Nucleus PURE: pET28a-ThrRS in pET28a protein expression plasmid. See
            https://nucleus.bnext.bio/ for more details..
ACCESSION   pET28a-ThrRS
VERSION     pET28a-ThrRS
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     rep_origin      77..665
                     /direction=RIGHT
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     promoter        77..105
                     /label="RNAII promoter"
                     /note="Color: #ffffff NCBI Feature Key: promoter NCBI Join
                     Type: join Transferred From: RNAII_promoter Transferred
                     Similarity: 100.00%"
     misc_RNA        113..665
                     /label="RNAII transcript (complementary strand)"
                     /note="RNAII transcript (complementary strand)"
     misc_RNA        complement(116..223)
                     /label="RNAI transcript"
                     /note="RNAI transcript"
     promoter        complement(230..259)
                     /label="RNA I promoter"
                     /note="Color: #ffffff NCBI Feature Key: promoter NCBI Join
                     Type: join Transferred From: RNA_I_promoter Transferred
                     Similarity: 100.00%"
     terminator      669..712
                     /label="rop/bom terminator"
     misc_binding    752..760
                     /label="dnaA binding site"
                     /bound_moiety="dnaA"
     misc_feature    785..848
                     /label="L-strand Y effector site"
                     /note="L-strand Y effector site"
     misc_feature    851..990
                     /label="bom"
                     /note="basis of mobility region from pBR322"
     misc_feature    1031..1187
                     /label="H-strand Y effector site"
                     /note="H-strand Y effector site"
     CDS             complement(1092..1283)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein"
                     /label="rop"
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     RBS             1284..1293
     misc_feature    complement(1359..1377)
                     /label="ROP calculated promoter"
     CDS             complement(2092..3174)
                     /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"
     promoter        complement(3175..3252)
                     /label="lacI promoter"
                     /note="Color: #ffffff NCBI Feature Key: promoter
                     Transferred From: lacI_promoter Transferred Similarity:
                     100.00%"
     promoter        3561..3579
                     /label="T7_promoter"
     protein_bind    3580..3604
                     /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             3613..3645
                     /label="UTR1"
                     /note="note: UTR1 NCBI Feature Key: misc_feature Color:
                     #cc0000 modified by: Andrey Transferred From: U4m - UTR1
                     Transferred Similarity: 100.00%"
     CDS             3649..3651
                     /codon_start=1
                     /label="Start"
                     /translation="M"
     CDS             3652..3669
                     /codon_start=1
                     /label="His6"
                     /translation="HHHHHH"
     CDS             3670..3684
                     /codon_start=1
                     /label="Linker"
                     /translation="GGGGS"
     CDS             3685..5610
                     /codon_start=1
                     /standard_name="ThrRS"
                     /label="ThrRS"
                     /translation="MPVITLPDGSQRHYDHAVSPMDVALDIGPGLAKACIAGRVNGELV
                     DACDLIENDAQLSIITAKDEEGLEIIRHSCAHLLGHAIKQLWPHTKMAIGPVIDNGFYY
                     DVDLDRTLTQEDVEALEKRMHELAEKNYDVIKKKVSWHEARETFANRGESYKVSILDEN
                     IAHDDKPGLYFHEEYVDMCRGPHVPNMRFCHHFKLMKTAGAYWRGDSNNKMLQRIYGTA
                     WADKKALNAYLQRLEEAAKRDHRKIGKQLDLYHMQEEAPGMVFWHNDGWTIFRELEVFV
                     RSKLKEYQYQEVKGPFMMDRVLWEKTGHWDNYKDAMFTTSSENREYCIKPMNCPGHVQI
                     FNQGLKSYRDLPLRMAEFGSCHRNEPSGSLHGLMRVRGFTQDDAHIFCTEEQIRDEVNG
                     CIRLVYDMYSTFGFEKIVVKLSTRPEKRIGSDEMWDRAEADLAVALEENNIPFEYQLGE
                     GAFYGPKIEFTLYDCLDRAWQCGTVQLDFSLPSRLSASYVGEDNERKVPVMIHRAILGS
                     MERFIGILTEEFAGFFPTWLAPVQVVIMNITDSQSEYVNELTQKLSNAGIRVKADLRNE
                     KIGFKIREHTLRRVPYMLVCGDKEVESGKVAVRTRRGKDLGSMDVNEVIEKLQQEIRSR
                     SLKQLEE"
     terminator      5684..5731
                     /label="T7 terminator"
                     /note="Color: #ffffff NCBI Feature Key: terminator
                     Transferred From: T7_terminator Transferred Similarity:
                     100.00%"
     rep_origin      5768..6223
                     /direction=RIGHT
                     /label="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     CDS             complement(6315..7130)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label="KanR"
                     /note="confers resistance to kanamycin"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
ORIGIN
        1 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
       61 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
      121 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
      181 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt
      241 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
      301 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
      361 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
      421 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
      481 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
      541 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
      601 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
      661 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
      721 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
      781 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
      841 cggaagagcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca
      901 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt atacactccg
      961 ctatcgctac gtgactgggt catggctgcg ccccgacacc cgccaacacc cgctgacgcg
     1021 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     1081 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa
     1141 agctcatcag cgtggtcgtg aagcgattca cagatgtctg cctgttcatc cgcgtccagc
     1201 tcgttgagtt tctccagaag cgttaatgtc tggcttctga taaagcgggc catgttaagg
     1261 gcggtttttt cctgtttggt cactgatgcc tccgtgtaag ggggatttct gttcatgggg
     1321 gtaatgatac cgatgaaacg agagaggatg ctcacgatac gggttactga tgatgaacat
     1381 gcccggttac tggaacgttg tgagggtaaa caactggcgg tatggatgcg gcgggaccag
     1441 agaaaaatca ctcagggtca atgccagcgc ttcgttaata cagatgtagg tgttccacag
     1501 ggtagccagc agcatcctgc gatgcagatc cggaacataa tggtgcaggg cgctgacttc
     1561 cgcgtttcca gactttacga aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc
     1621 gcagacgttt tgcagcagca gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc
     1681 taaccagtaa ggcaaccccg ccagcctagc cgggtcctca acgacaggag cacgatcatg
     1741 cgcacccgtg gggccgccat gccggcgata atggcctgct tctcgccgaa acgtttggtg
     1801 gcgggaccag tgacgaaggc ttgagcgagg gcgtgcaaga ttccgaatac cgcaagcgac
     1861 aggccgatca tcgtcgcgct ccagcgaaag cggtcctcgc cgaaaatgac ccagagcgct
     1921 gccggcacct gtcctacgag ttgcatgata aagaagacag tcataagtgc ggcgacgata
     1981 gtcatgcccc gcgcccaccg gaaggagctg actgggttga aggctctcaa gggcatcggt
     2041 cgagatcccg gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg ctcactgccc
     2101 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     2161 agaggcggtt tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg agacgggcaa
     2221 cagctgattg cccttcaccg cctggccctg agagagttgc agcaagcggt ccacgctggt
     2281 ttgccccagc aggcgaaaat cctgtttgat ggtggttaac ggcgggatat aacatgagct
     2341 gtcttcggta tcgtcgtatc ccactaccga gatatccgca ccaacgcgca gcccggactc
     2401 ggtaatggcg cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg
     2461 aacgatgccc tcattcagca tttgcatggt ttgttgaaaa ccggacatgg cactccagtc
     2521 gccttcccgt tccgctatcg gctgaatttg attgcgagtg agatatttat gccagccagc
     2581 cagacgcaga cgcgccgaga cagaacttaa tgggcccgct aacagcgcga tttgctggtg
     2641 acccaatgcg accagatgct ccacgcccag tcgcgtaccg tcttcatggg agaaaataat
     2701 actgttgatg ggtgtctggt cagagacatc aagaaataac gccggaacat tagtgcaggc
     2761 agcttccaca gcaatggcat cctggtcatc cagcggatag ttaatgatca gcccactgac
     2821 gcgttgcgcg agaagattgt gcaccgccgc tttacaggct tcgacgccgc ttcgttctac
     2881 catcgacacc accacgctgg cacccagttg atcggcgcga gatttaatcg ccgcgacaat
     2941 ttgcgacggc gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca acgactgttt
     3001 gcccgccagt tgttgtgcca cgcggttggg aatgtaattc agctccgcca tcgccgcttc
     3061 cactttttcc cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt
     3121 ctgataagag acaccggcat actctgcgac atcgtataac gttactggtt tcacattcac
     3181 caccctgaat tgactctctt ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca
     3241 ttcgatggtg tccgggatct cgacgctctc ccttatgcga ctcctgcatt aggaagcagc
     3301 ccagtagtag gttgaggccg ttgagcaccg ccgccgcaag gaatggtgca tgcaaggaga
     3361 tggcgcccaa cagtcccccg gccacggggc ctgccaccat acccacgccg aaacaagcgc
     3421 tcatgagccc gaagtggcga gcccgatctt ccccatcggt gatgtcggcg atataggcgc
     3481 cagcaaccgc acctgtggcg ccggtgatgc cggccacgat gcgtccggcg tagaggatcg
     3541 agatctcgat cccgcgaaat taatacgact cactataggg gaattgtgag cggataacaa
     3601 ttcccctcta gaaataattt tgtttaactt taagaaggag atataccaat gcaccatcac
     3661 caccatcatg gaggcggtgg atctatgccg gttatcaccc tgccagatgg ttctcagcgt
     3721 cattatgatc acgctgtcag tcctatggat gtggcacttg acatcggccc gggcttggcc
     3781 aaggcgtgta ttgcaggacg tgttaatggt gaactggtag atgcttgtga tctgatcgag
     3841 aacgatgccc aactgagcat catcactgct aaagatgaag aaggtcttga aatcattcgc
     3901 catagctgtg cccatctgtt gggccacgca attaaacagc tgtggccgca cacaaaaatg
     3961 gcaattggcc ctgtcatcga taatgggttt tattatgacg tggacctgga ccggacgctg
     4021 acacaagaag atgtagaggc attggaaaaa cggatgcatg aactggccga aaaaaattat
     4081 gacgttatta agaaaaaagt ctcctggcat gaggcgcgcg aaacatttgc gaaccgcggc
     4141 gagagctata aagtctctat tctggacgag aatattgccc atgatgataa gccgggtctg
     4201 tatttccatg aggaatatgt ggacatgtgc cggggtcctc acgtgccgaa catgcgtttc
     4261 tgccaccact ttaaattgat gaaaaccgcc ggtgcatatt ggcgcggtga ttccaataac
     4321 aagatgctgc agcgcattta cggtaccgcc tgggcggata aaaaagccct gaacgcgtac
     4381 ttacagcgtc tggaagaggc cgctaaacgg gatcatcgta aaattggtaa gcagctggat
     4441 ttatatcata tgcaggaaga ggcgcctggg atggtctttt ggcataacga tggctggacc
     4501 attttccgtg aactggaagt ctttgtacgc agtaaattaa aagaatacca gtatcaggag
     4561 gttaaaggac ctttcatgat ggatcgggtt ctgtgggaaa aaaccggcca ttgggataac
     4621 tataaggatg ccatgttcac gacatcgtcg gaaaatcggg aatactgtat taaacctatg
     4681 aactgtccgg gccatgtgca gatttttaac caaggcctta aaagttatcg tgatctgcca
     4741 ttacgcatgg ccgaatttgg gtcgtgccat cgtaatgagc catccggcag cttgcacggt
     4801 ttgatgcgcg ttcgcggatt tactcaggat gatgctcata ttttttgcac cgaggagcag
     4861 atccgtgacg aggttaatgg ttgcatccgc cttgtctacg atatgtactc aacgtttggc
     4921 ttcgagaaga tcgtggtgaa gctgtcgaca cgtccggaaa aacgtattgg ctccgacgaa
     4981 atgtgggatc gtgcggaggc agaccttgcg gtggcattgg aggagaataa catccccttc
     5041 gagtaccagc tgggtgaggg tgctttttac ggcccgaaaa ttgaatttac gttatacgat
     5101 tgcttggatc gggcgtggca gtgcgggaca gttcaattgg atttctcatt gccatcccgc
     5161 ctgagcgcga gctacgtggg tgaggataat gaacgcaaag ttccggtaat gatccatcgg
     5221 gccattctgg gcagtatgga acgttttatc ggcatcttaa cggaagaatt tgctggtttt
     5281 ttcccgactt ggctggcgcc cgttcaggtc gtgatcatga acattaccga ttctcaatcg
     5341 gaatatgtga atgaactgac tcagaagctg tcaaatgctg gtattcgtgt caaggccgat
     5401 cttcgcaatg agaaaatcgg cttcaaaatc cgcgaacaca cgcttcgccg cgtaccatat
     5461 atgctggttt gtggtgacaa agaagttgaa tctggtaaag ttgctgtgcg gacgcgtcgt
     5521 ggcaaagatt tggggtcgat ggacgtcaac gaggtgattg aaaaactgca acaggaaatc
     5581 cgctcacgtt ccctgaaaca gctggaggag tgaagagctt gatccggctg ctaacaaagc
     5641 ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg
     5701 ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc
     5761 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     5821 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     5881 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     5941 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     6001 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     6061 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     6121 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     6181 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg
     6241 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     6301 tcatgaatta attcttagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata
     6361 tcaggattat caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca
     6421 ccgaggcagt tccataggat ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca
     6481 acatcaatac aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca
     6541 ccatgagtga cgactgaatc cggtgagaat ggcaaaagtt tatgcatttc tttccagact
     6601 tgttcaacag gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta
     6661 ttcattcgtg attgcgcctg agcgagacga aatacgcgat cgctgttaaa aggacaatta
     6721 caaacaggaa tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca
     6781 cctgaatcag gatattcttc taatacctgg aatgctgttt tcccggggat cgcagtggtg
     6841 agtaaccatg catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat
     6901 tccgtcagcc agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg
     6961 ccatgtttca gaaacaactc tggcgcatcg ggcttcccat acaatcgata gattgtcgca
     7021 cctgattgcc cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg
     7081 gaatttaatc gcggcctaga gcaagacgtt tcccgttgaa tatggctcat aacacccctt
     7141 gtattactgt ttatgtaagc agacagtttt attgt
//
