LOCUS       sequence-83360-e        7696 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a plant arginyl-tRNA synthetase in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7696)
  AUTHORS   Igloi GL, Schiefermayr E
  TITLE     Amino acid discrimination by arginyl-tRNA synthetases as revealed by
            an examination of natural specificity variants.
  JOURNAL   FEBS J. 2009 Mar;276(5):1307-18. doi: 
            10.1111/j.1742-4658.2009.06866.x.
  PUBMED    19187230
REFERENCE   2  (bases 1 to 7696)
  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: 
            "10.1111/j.1742-4658.2009.06866"; journalName: "FEBS J"; date: 
            "2009-03"; volume: "276"; issue: "5"; pages: "1307-18"
COMMENT     Sequence Label: pET-Gm1
FEATURES             Location/Qualifiers
     source          1..7696
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(2015..2029)
                     /codon_start=1
                     /product="enterokinase recognition and cleavage site"
                     /label=enterokinase site
                     /translation="DDDDK"
     CDS             complement(2045..2089)
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             complement(2096..2113)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(2123..2140)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(2162..2488)
                     /codon_start=1
                     /gene="trxA"
                     /product="E. coli thioredoxin"
                     /label=TrxA
                     /translation="MSDKIIHLTDDSFDTDVLKADGAILVDFWAEWCGPCKMIAPILDE
                     IADEYQGKLTVAKLNIDQNPGTAPKYGIRGIPTLLLFKNGEVAATKVGALSKGQLKEFL
                     DANLA"
     RBS             2496..2518
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    2533..2557
                     /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(2557..2576)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2558..2576)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2664..2683)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        2889..2966
                     /gene="lacI"
                     /label=lacI promoter
     CDS             2967..4049
                     /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(2986..3005)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4858..5049
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(5043..5065)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    5151..5293
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     5206..5225
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5308..5325)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5479..6067)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5559..5578)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6238..7029,7030..7098))
                     /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     6861..6880
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7099..7203)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(7230..7685)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7367..7388)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7579..7598
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagcttgt
      181 cgacggagct cgaattcgga tccgatatca gccatggtca tagcctgtat acaggttcaa
      241 ttccaaggag ataaaagcat tgtctcatca cagtcactgt tgcttcacat agcaagagtc
      301 tactagtctc ctcaggcgac cccacaacct ggcaattagt ataaaatttt tttgtaaaga
      361 tttctgtcaa attatagagg tattcacaca gcacattggg caacaaattg gtcaacgact
      421 cctcaaaaac ctcaggaaat tgtatcaaat gaagccccaa tgcacgttca tcttcatgat
      481 tcgacactat attcccattt ttctttattt cttccatatc tttaccagat ttcctcatga
      541 tagaacagat cctagcatgc gcatactgca aataaacagc agtatttccc ttatcattaa
      601 gcatctgatc aaagctgaat gtgtaattcg ttaatctgtt gatcctcaag tcagcatact
      661 taacagctcc gtaaccaact gcttctgctg tcttttcgat ctcttcctca gaccaatctt
      721 taacagcatc acgctcgagg agggaagctt tacaacgcct tttggcttca tcaagcaact
      781 caaccaatct aacagtttca ctgctgcgtg tccgaaatcg ttttccatct tccccaagaa
      841 caagaccaaa accaatatga gtacatttgg ggaactcatt ttcatgattt ggcagccatc
      901 ctgcacgcct aaaggcctta aaaaccatat caaagtgttg ccactgccca acatctgtaa
      961 cgtatatatt ccactcaaca ttttccacat ttagacgatt ccaaagtgct gctagatcag
     1021 ttgaagaata gttgtagcca ccatctcttt tcacagcaat aattggtata tttacaccct
     1081 caacaaatat cacacgagca ccatcatctt cttcaatcag tcctaaatta tccaatctct
     1141 ccaaagtttg agggattaaa tcattatagt agctctctcc ccttgcttcc aatcgaactc
     1201 caagacgttg atagaccttt tcaaattcag cctgactaat atcacaaatt tgctgccatg
     1261 ccctgtgata cttttcttct ccactctgga gccggaccac tgactgttgt gccctcaact
     1321 taaattcagg atcattatca aacctcactt tggaggcttt atagaatgcc tgaagatccc
     1381 caatagctgc ttcattaaaa tcttctgggt ttggataagt gtcaaagagg tgtgctatta
     1441 gcatcccaaa ctgagtcccc cagtcaccaa gatgatttct gcggataaga gtttccacac
     1501 gtgaaaattc aaacatgcga gctaatgtgt ccccaataat ggtagatctc aggtgtccaa
     1561 catgcatttc ctttgctatg ttaggtgagg aaaaatcaac cagaactgtc ttcacgggaa
     1621 gtcgaggtgc ccacttatca ataccatcaa tcagcaacct ctctaacctc tgcgctatcc
     1681 acttcttcga caagacaata ttcacaaacc caggaccagc tacagagcat gattctatca
     1741 tttcagactg cggaagatgg ttaagtatgg cctgtccaac tgcctgggga cccctgaatc
     1801 ctgtctgctt ccctttcatc ttagcaaata gacccattgc attattacat tgataatcag
     1861 cgaatttaat accagctttg ttggcagcgc aaggatcaat agaaggcaca acatcgggtt
     1921 catcaggcac ggttatccgg agagactcct caaactcctt cgccaactgc ttcttaacac
     1981 tagctggaca atatacttca acatacgcca tggccttgtc gtcgtcgtcg gtacccagat
     2041 ctgggctgtc catgtgctgg cgttcgaatt tagcagcagc ggtttctttc ataccagaac
     2101 cgcgtggcac cagaccagaa gaatgatgat gatgatggtg catatggcca gaaccagaac
     2161 cggccaggtt agcgtcgagg aactctttca actgaccttt agacagtgca cccactttgg
     2221 ttgccgccac ttcaccgttt ttgaacagca gcagagtcgg gataccacgg atgccatatt
     2281 tcggcgcagt gccagggttt tgatcgatgt tcagttttgc aacggtcagt ttgccctgat
     2341 attcgtcagc gatttcatcc agaatcgggg cgatcatttt gcacggaccg caccactctg
     2401 cccagaaatc gacgaggatc gccccgtccg ctttgagtac atccgtgtca aaactgtcgt
     2461 cagtcaggtg aataatttta tcgctcatat gtatatctcc ttcttaaagt taaacaaaat
     2521 tatttctaga ggggaattgt tatccgctca caattcccct atagtgagtc gtattaattt
     2581 cgcgggatcg agatcgatct cgatcctcta cgccggacgc atcgtggccg gcatcaccgg
     2641 cgccacaggt gcggttgctg gcgcctatat cgccgacatc accgatgggg aagatcgggc
     2701 tcgccacttc gggctcatga gcgcttgttt cggcgtgggt atggtggcag gccccgtggc
     2761 cgggggactg ttgggcgcca tctccttgca tgcaccattc cttgcggcgg cggtgctcaa
     2821 cggcctcaac ctactactgg gctgcttcct aatgcaggag tcgcataagg gagagcgtcg
     2881 agatcccgga caccatcgaa tggcgcaaaa cctttcgcgg tatggcatga tagcgcccgg
     2941 aagagagtca attcagggtg gtgaatgtga aaccagtaac gttatacgat gtcgcagagt
     3001 atgccggtgt ctcttatcag accgtttccc gcgtggtgaa ccaggccagc cacgtttctg
     3061 cgaaaacgcg ggaaaaagtg gaagcggcga tggcggagct gaattacatt cccaaccgcg
     3121 tggcacaaca actggcgggc aaacagtcgt tgctgattgg cgttgccacc tccagtctgg
     3181 ccctgcacgc gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat caactgggtg
     3241 ccagcgtggt ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc
     3301 acaatcttct cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg gatgaccagg
     3361 atgccattgc tgtggaagct gcctgcacta atgttccggc gttatttctt gatgtctctg
     3421 accagacacc catcaacagt attattttct cccatgaaga cggtacgcga ctgggcgtgg
     3481 agcatctggt cgcattgggt caccagcaaa tcgcgctgtt agcgggccca ttaagttctg
     3541 tctcggcgcg tctgcgtctg gctggctggc ataaatatct cactcgcaat caaattcagc
     3601 cgatagcgga acgggaaggc gactggagtg ccatgtccgg ttttcaacaa accatgcaaa
     3661 tgctgaatga gggcatcgtt cccactgcga tgctggttgc caacgatcag atggcgctgg
     3721 gcgcaatgcg cgccattacc gagtccgggc tgcgcgttgg tgcggacatc tcggtagtgg
     3781 gatacgacga taccgaagac agctcatgtt atatcccgcc gttaaccacc atcaaacagg
     3841 attttcgcct gctggggcaa accagcgtgg accgcttgct gcaactctct cagggccagg
     3901 cggtgaaggg caatcagctg ttgcccgtct cactggtgaa aagaaaaacc accctggcgc
     3961 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
     4021 aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtaag ttagctcact
     4081 cattaggcac cgggatctcg accgatgccc ttgagagcct tcaacccagt cagctccttc
     4141 cggtgggcgc ggggcatgac tatcgtcgcc gcacttatga ctgtcttctt tatcatgcaa
     4201 ctcgtaggac aggtgccggc agcgctctgg gtcattttcg gcgaggaccg ctttcgctgg
     4261 agcgcgacga tgatcggcct gtcgcttgcg gtattcggaa tcttgcacgc cctcgctcaa
     4321 gccttcgtca ctggtcccgc caccaaacgt ttcggcgaga agcaggccat tatcgccggc
     4381 atggcggccc cacgggtgcg catgatcgtg ctcctgtcgt tgaggacccg gctaggctgg
     4441 cggggttgcc ttactggtta gcagaatgaa tcaccgatac gcgagcgaac gtgaagcgac
     4501 tgctgctgca aaacgtctgc gacctgagca acaacatgaa tggtcttcgg tttccgtgtt
     4561 tcgtaaagtc tggaaacgcg gaagtcagcg ccctgcacca ttatgttccg gatctgcatc
     4621 gcaggatgct gctggctacc ctgtggaaca cctacatctg tattaacgaa gcgctggcat
     4681 tgaccctgag tgatttttct ctggtcccgc cgcatccata ccgccagttg tttaccctca
     4741 caacgttcca gtaaccgggc atgttcatca tcagtaaccc gtatcgtgag catcctctct
     4801 cgtttcatcg gtatcattac ccccatgaac agaaatcccc cttacacgga ggcatcagtg
     4861 accaaacagg aaaaaaccgc ccttaacatg gcccgcttta tcagaagcca gacattaacg
     4921 cttctggaga aactcaacga gctggacgcg gatgaacagg cagacatctg tgaatcgctt
     4981 cacgaccacg ctgatgagct ttaccgcagc tgcctcgcgc gtttcggtga tgacggtgaa
     5041 aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg
     5101 agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg
     5161 acccagtcac gtagcgatag cggagtgtat actggcttaa ctatgcggca tcagagcaga
     5221 ttgtactgag agtgcaccat atatgcggtg tgaaataccg cacagatgcg taaggagaaa
     5281 ataccgcatc aggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     5341 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     5401 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     5461 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     5521 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     5581 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     5641 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     5701 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     5761 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     5821 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     5881 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     5941 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     6001 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     6061 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     6121 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     6181 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     6241 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     6301 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     6361 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     6421 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     6481 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     6541 tgttgccatt gctgcaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     6601 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     6661 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     6721 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     6781 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     6841 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     6901 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     6961 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     7021 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     7081 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     7141 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     7201 gcgcacattt ccccgaaaag tgccacctga aattgtaaac gttaatattt tgttaaaatt
     7261 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     7321 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     7381 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     7441 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     7501 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     7561 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     7621 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     7681 cgcgtcccat tcgcca
//
