LOCUS       sequence-183278-        6904 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  express Methanosarcina barkeri pyrrolysyl-tRNA synthetase/tRNA pair 
            in E. coli to incorporate photocrosslinkers DiZPK, DiZSeK, or 
            DiZHSeC into an in-frame amber codon site on a protein of interest.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6904)
  AUTHORS   Zhang M, Lin S, Song X, Liu J, Fu Y, Ge X, Fu X, Chang Z, Chen PR
  TITLE     A genetically incorporated crosslinker reveals chaperone cooperation
            in acid resistance.
  JOURNAL   Nat Chem Biol. 2011 Sep 4;7(10):671-7. doi: 10.1038/nchembio.644.
  PUBMED    21892184
REFERENCE   2  (bases 1 to 6904)
  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: "10.1038/nchembio"; 
            journalName: "Nat Chem Biol"; date: "2011-09-4- 4"; volume: "7"; 
            issue: "10"; pages: "671-7"
COMMENT     Sequence Label: pSupAR-MbPylRS(DiZPK)
FEATURES             Location/Qualifiers
     source          1..6904
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        20..304
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     227..246
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     primer_bind     310..329
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        310..328
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             331..353
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(1668..1685)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1838..1884
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(4040..4699)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     4614..4633
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(4700..4802)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(5194..5211)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5328..5873)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(5408..5427)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(6019..6897)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPSDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
ORIGIN
        1 atactcccgc cattcagaga agaaaccaat tgtccatatt gcatcagaca ttgccgtcac
       61 tgcgtctttt actggctctt ctcgctaacc aaaccggtaa ccccgcttat taaaagcatt
      121 ctgtaacaaa gcgggaccaa agccatgaca aaaacgcgta acaaaagtgt ctataatcac
      181 ggcagaaaag tccacattga ttatttgcac ggcgtcacac tttgctatgc catagcattt
      241 ttatccataa gattagcgga tcctacctga cgctttttat cgcaactctc tactgtttct
      301 ccatacccgt aatacgactc actatagggt tttgtttaac tttaagaagg agatatactt
      361 acccatggat aaaaaaccat tagatgtttt aatatctgcg accgggctct ggatgtccag
      421 gactggcacg ctccacaaaa tcaagcacca tgaggtctca agaagtaaaa tatacattga
      481 aatggcgtgt ggagaccatc ttgttgtgaa taattccagg agttgtagaa cagccagagc
      541 attcagacat cataagtaca gaaaaacctg caaacgatgt agggtttcgg acgaggatat
      601 caataatttt ctcacaagat caaccgaaag caaaaacagt gtgaaagtta gggtagtttc
      661 tgctccaaag gtcaaaaaag ctatgccgaa atcagtttca agggctccga agcctctgga
      721 aaattctgtt tctgcaaagg catcaacgaa cacatccaga tctgtacctt cgcctgcaaa
      781 atcaactcca aattcgtctg ttcccgcatc ggctcctgct ccttcactta caagaagcca
      841 gcttgatagg gttgaggctc tcttaagtcc agaggataaa atttctctaa atatggcaaa
      901 gcctttcagg gaacttgagc ctgaacttgt gacaagaaga aaaaacgatt ttcagcggct
      961 ctataccaat gatagagaag actacctcgg taaactcgaa cgtgatatta cgaaattttt
     1021 cgtagaccgg ggttttctgg agataaagtc tcctatcctt attccggcgg aatacgtgga
     1081 gagaatgggt attaataatg atactgaact ttcaaaacag atcttccggg tggataaaaa
     1141 tctctgcttg aggccaatgc ttgccccgac tctgtataac tatgcgcgaa aactcgatag
     1201 gattttacca ggcccaataa aaattttcga agtcggacct tgttaccgga aagagtctga
     1261 cggcaaagag cacctggaag aatttactat ggtgaacttc agtcagatgg gttcgggatg
     1321 tactcgggaa aatcttgaag ctctcatcaa agagtttctg gactatctgg aaatcgactt
     1381 cgaaatcgta ggagattcct gtatggtctt tggggatact cttgatataa tgcacgggga
     1441 cctggagctt tcttcggcag tcgtcgggcc agtttctctt gatagagaat ggggtattga
     1501 caaaccatgg ataggtgcag gttttggtct tgaacgcttg ctcaaggtta tgcacggctt
     1561 taaaaacatt aagagggcat caaggtccga atcttactat aatgggattt caaccaatct
     1621 ataagtctcc agcttggctg ttttggcgga tgagagaaga ttttcagcct gatacagatt
     1681 aaatcagaac gcagaagcgg tctgataaaa cagaatttgc ctggcggcag tagcgcggtg
     1741 gtcccacctg accccatgcc gaactcagaa gtgaaacgcc gtagcgccga tggtagtgtg
     1801 gggtctcccc atgcgagagt agggaactgc caggcatcaa ataaaacgaa aggctcagtc
     1861 gaaagactgg gcctttcgtt ttatctgttg tttgtgagct cccggtcatc aatcatcccc
     1921 ataatccttg ttagattatc aattttaaaa aactaacagt tgtcagcctg tcccgcttta
     1981 atatcatacg ccgttatacg ttgtttacgc tttgaggaat cccatatgga taaaaagcct
     2041 ctgaacactc tgatttctgc gaccggtctg tggatgtccc gcaccggcac catccacaaa
     2101 atcaaacacc atgaagttag ccgttccaaa atctacattg aaatggcttg cggcgatcac
     2161 ctggttgtca acaactcccg ttcttctcgt accgctcgcg cactgcgcca ccacaaatat
     2221 cgcaaaacct gcaaacgttg ccgtgttagc gatgaagatc tgaacaaatt cctgaccaaa
     2281 gctaacgagg atcagacctc cgtaaaagtg aaggtagtaa gcgctccgac ccgtactaaa
     2341 aaggctatgc caaaaagcgt ggcccgtgcc ccgaaacctc tggaaaacac cgaggcggct
     2401 caggctcaac catccggttc taaattttct ccggcgatcc cagtgtccac ccaagaatct
     2461 gtttccgtac cagcaagcgt gtctaccagc attagcagca tttctaccgg tgctaccgct
     2521 tctgcgctgg taaaaggtaa cactaacccg attactagca tgtctgcacc ggtacaggca
     2581 agcgccccag ctctgactaa atcccagacg gaccgtctgg aggtgctgct gaacccaaag
     2641 gatgaaatct ctctgaacag cggcaagcct ttccgtgagc tggaaagcga gctgctgtct
     2701 cgtcgtaaaa aggatctgca acagatctac gctgaggaac gcgagaacta tctgggtaag
     2761 ctggagcgcg aaattactcg cttcttcgtg gatcgcggtt tcctggagat caaatctccg
     2821 attctgattc cgctggaata cattgaacgt atgggcatcg ataatgatac cgaactgtct
     2881 aaacagatct tccgtgtgga taaaaacttc tgtctgcgtc cgatgctggc cccgaacctg
     2941 tacaactatc tgcgtaaact ggaccgtgcc ctgccggacc cgatcaaaat tttcgagatc
     3001 ggtccttgct accgtaaaga gtccgacggt aaagagcacc tggaagagtt taccatgctg
     3061 aacttctgcc agatgggtag cggttgcacg cgtgaaaacc tggaatccat tatcaccgac
     3121 ttcctgaatc acctgggtat cgatttcaaa attgttggtg acagctgtat ggtgtacggc
     3181 gatacgctgg atgttatgca cggcgatctg gagctgtctt ccgcagtagt gggcccaatc
     3241 ccgctggatc gtgagtgggg tatcgacaaa ccttggatcg gtgcgggttt tggtctggag
     3301 cgtctgctga aagtaaaaca cgacttcaag aacatcaaac gtgctgcacg ttccgagtcc
     3361 tattacaatg gtatttctac taacctgtaa ctgcagtttc aaacgctaaa ttgcctgatg
     3421 cgctacgctt atcaggccta catgatctct gcaatatatt gagtttgcgt gcttttgtag
     3481 gccggataag gcgttcacgc cgcatccggc aagaaacagc aaacaatcca aaacgccgcg
     3541 ttcagcggcg ttttttctgc ttttcttcgc gaattaattc cgcttcgcaa catgtgagca
     3601 ccggtttatt gactaccgga agcagtgtga ccgtgtgctt ctcaaatgcc tgaggccagt
     3661 ttgctcaggc tctccccgtg gaggtaataa ttgacgatat gatcagtgca cggctaacta
     3721 agcggcctgc tgactttctc gccgatcaaa aggcattttg ctattaaggg attgacgagg
     3781 gcgtatctgc gcagtaagat gcgccccgca ttggaaacct gatcatgtag atcgaatgga
     3841 ctctaaatcc gttcagccgg gttagattcc cggggtttcc gccaaattcg aaaagcctgc
     3901 tcaacgagca ggcttttttg catctcgagc agctcagggt cgaatttgct ttcgaatttc
     3961 tgccattcat ccgcttatta tcacttattc aggcgtagca accaggcgtt taagggcacc
     4021 aataactgcc ttaaaaaaat tacgccccgc cctgccactc atcgcagtac tgttgtaatt
     4081 cattaagcat tctgccgaca tggaagccat cacaaacggc atgatgaacc tgaatcgcca
     4141 gcggcatcag caccttgtcg ccttgcgtat aatatttgcc catggtgaaa acgggggcga
     4201 agaagttgtc catattggcc acgtttaaat caaaactggt gaaactcacc cagggattgg
     4261 ctgagacgaa aaacatattc tcaataaacc ctttagggaa ataggccagg ttttcaccgt
     4321 aacacgccac atcttgcgaa tatatgtgta gaaactgccg gaaatcgtcg tggtattcac
     4381 tccagagcga tgaaaacgtt tcagtttgct catggaaaac ggtgtaacaa gggtgaacac
     4441 tatcccatat caccagctca ccgtctttca ttgccatacg gaattccgga tgagcattca
     4501 tcaggcgggc aagaatgtga ataaaggccg gataaaactt gtgcttattt ttctttacgg
     4561 tctttaaaaa ggccgtaata tccagctgaa cggtctggtt ataggtacat tgagcaactg
     4621 actgaaatgc ctcaaaatgt tctttacgat gccattggga tatatcaacg gtggtatatc
     4681 cagtgatttt tttctccatt ttagcttcct tagctcctga aaatctcgat aactcaaaaa
     4741 atacgcccgg tagtgatctt atttcattat ggtgaaagtt ggaacctctt acgtgccgat
     4801 caacgtctca ttttcgccaa aagttggccc agggcttccc ggtatcaaca gggacaccag
     4861 gatttattta ttctgcgaag tgatcttccg tcacaggtat ttattcggcg caaagtgcgt
     4921 cgggtgatgc tgccaactta ctgatttagt gtatgatggt gtttttgagg tgctccagtg
     4981 gcttctgttt ctatcagctg tccctcctgt tcagctactg acggggtggt gcgtaacggc
     5041 aaaagcaccg ccggacatca gcgctagcgg agtgtatact ggcttactat gttggcactg
     5101 atgagggtgt cagtgaagtg cttcatgtgg caggagaaaa aaggctgcac cggtgcgtca
     5161 gcagaatatg tgatacagga tatattccgc ttcctcgctc actgactcgc tacgctcggt
     5221 cgttcgactg cggcgagcgg aaatggctta cgaacggggc ggagatttcc tggaagatgc
     5281 caggaagata cttaacaggg aagtgagagg gccgcggcaa agccgttttt ccataggctc
     5341 cgcccccctg acaagcatca cgaaatctga cgctcaaatc agtggtggcg aaacccgaca
     5401 ggactataaa gataccaggc gtttccccct ggcggctccc tcgtgcgctc tcctgttcct
     5461 gcctttcggt ttaccggtgt cattccgctg ttatggccgc gtttgtctca ttccacgcct
     5521 gacactcagt tccgggtagg cagttcgctc caagctggac tgtatgcacg aaccccccgt
     5581 tcagtccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggaaagaca
     5641 tgcaaaagca ccactggcag cagccactgg taattgattt agaggagtta gtcttgaagt
     5701 catgcgccgg ttaaggctaa actgaaagga caagttttgg tgactgcgct cctccaagcc
     5761 agttacctcg gttcaaagag ttggtagctc agagaacctt cgaaaaaccg ccctgcaagg
     5821 cggttttttc gttttcagag caagagatta cgcgcagacc aaaacgatct caagaagatc
     5881 atcttattaa tcagataaaa tatttctaga tttcagtgca atttatctct tcaaatgtag
     5941 cacctgaagt cagccccata cgatataagt tgtaattctc atgtttgaca gcttatcatc
     6001 gataagcttg gtacccaatt atgacaactt gacggctaca tcattcactt tttcttcaca
     6061 accggcacgg aactcgctcg ggctggcccc ggtgcatttt ttaaataccc gcgagaaata
     6121 gagttgatcg tcaaaaccaa cattgcgacc gacggtggcg ataggcatcc gggtggtgct
     6181 caaaagcagc ttcgcctggc tgatacgttg gtcctcgcgc cagcttaaga cgctaatccc
     6241 taactgctgg cggaaaagat gtgacagacg cgacggcgac aagcaaacat gctgtgcgac
     6301 gctggcgata tcaaaattgc tgtctgccag gtgatcgctg atgtactgac aagcctcgcg
     6361 tacccgatta tccatcggtg gatggagcga ctcgttaatc gcttccatgc gccgcagtaa
     6421 caattgctca agcagattta tcgccagcag ctccgaatag cgcccttccc cttgcccggc
     6481 gttaatgatt tgcccaaaca ggtcgctgaa atgcggctgg tgcgcttcat ccgggcgaaa
     6541 gaaccccgta ttggcaaata ttgacggcca gttaagccat tcatgccagt aggcgcgcgg
     6601 acgaaagtaa acccactggt gataccattc gcgagcctcc ggatgacgac cgtagtgatg
     6661 aatctctcct ggcgggaaca gcaaaatatc actcggtcgg caaacaaatt ctcgtccctg
     6721 atttttcacc accccctgac cgcgaatggt gagattgaga atataacctt tcattcccag
     6781 cggtcggtcg ataaaaaaat cgagataacc gttggcctca atcggcgtta aacccgccac
     6841 cagatgggca ttaaacgagt atcccggcag caggggatca ttttgcgctt cagccatact
     6901 tttc
//
