LOCUS       pAS_4xh7SKPylT_EF1_NES- 9449 bp ds-DNA     circular     06-OCT-2021
DEFINITION  .
FEATURES             Location/Qualifiers
     misc_feature    complement(1..105)
                     /label="bla Promoter"
     misc_feature    complement(131..586)
                     /label="f1 bacteriophage origin of replication; arrowindicates direction of (+) strand synthesis"
     misc_feature    705..727
                     /label="M13 uni (-43) (mismatches: 0)"
     misc_feature    727..744
                     /label="M13 uni (-21) (mismatches: 0)"
     misc_feature    728..744
                     /label="common sequencing primer, one of multiple similarvariants"
     misc_feature    871..1110
                     /label="PB3'"
     misc_feature    1116..1344
                     /label="Core Insulator"
     misc_feature    1348..1349
                     /label="15mer SpeI_up"
     misc_feature    complement(1366..1370)
                     /label="U6 terminator"
     misc_feature    complement(1382..1450)
                     /label="PylT"
     promoter        complement(1451..1693)
                     /label="h7SK promoter"
     misc_feature    complement(1695..1699)
                     /label="U6 terminator"
     misc_feature    complement(1711..1779)
                     /label="PylT"
     promoter        complement(1780..2022)
                     /label="h7SK promoter"
     misc_feature    complement(2024..2028)
                     /label="U6 terminator"
     misc_feature    complement(2040..2108)
                     /label="PylT"
     promoter        complement(2109..2351)
                     /label="h7SK promoter"
     misc_feature    complement(2353..2357)
                     /label="U6 terminator"
     misc_feature    complement(2369..2437)
                     /label="PylT"
     promoter        complement(2438..2680)
                     /label="h7SK promoter"
     promoter        2696..3248
                     /label="EF1"
     CDS             3258..3284
                     /label="Translation 3258-3284"
     CDS             3258..3323
                     /label="FLAG-NES-MmaPylRS AF (Y271A, Y349F)"
     CDS             3261..3284
                     /label="FLAG"
     CDS             3285..3323
                     /label="Translation 3285-3323"
     CDS             3324..4685
                     /label="PylRS Y306A, Y384F"
     CDS             3324..4685
                     /label="Translation 3324-4685"
     misc_feature    4707..5293
                     /label="internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)"
     CDS             5306..5905
                     /label="pac from Streptomyces alboniger CDS"
     CDS             5306..5905
                     /label="Translation 5306-5905"
     misc_feature    6020..6151
                     /label="SV40 EEL poly(A)"
     misc_feature    6111..6130
                     /label="pCEP-Reverse (mismatches: 0)"
     misc_feature    complement(6398..6626)
                     /label="Core Insulator"
     misc_feature    6632..6939
                     /label="PB5'"
     CDS             7127..7498
                     /label="Length: 372;Found at strand: positive;Start codon:ATG"
     misc_feature    complement(7427..7444)
                     /label="M13 rev (-29) (mismatches: 0)"
     misc_feature    complement(7428..7444)
                     /label="common sequencing primer, one of multiple similarvariants"
     misc_feature    7452..7468
                     /label="The lac repressor binds to the lac operator toinhibit transcription in E. coli. This inhibition canberelieved by adding lactoseorisopropyl-beta-D-thiogalactopyranoside (IPTG)."
     misc_feature    complement(7476..7506)
                     /label="promoter for the E. coli lac operon"
     misc_feature    complement(7486..7511)
                     /label="pEX-Rev (mismatches: 0)"
     misc_feature    7521..7542
                     /label="CAP binding activates transcription in the presenceof cAMP."
     misc_feature    complement(7830..8418)
                     /label="high-copy-number ColE1/pMB1/pBR322/pUC origin ofreplication"
     CDS             complement(8589..9449)
                     /label="bla CDS"
ORIGIN
        1 ACTCTTCCTT TTTCAATATT ATTGAAGCAT TTATCAGGGT TATTGTCTCA TGAGCGGATA
       61 CATATTTGAA TGTATTTAGA AAAATAAACA AATAGGGGTT CCGCGCACAT TTCCCCGAAA
      121 AGTGCCACCT AAATTGTAAG CGTTAATATT TTGTTAAAAT TCGCGTTAAA TTTTTGTTAA
      181 ATCAGCTCAT TTTTTAACCA ATAGGCCGAA ATCGGCAAAA TCCCTTATAA ATCAAAAGAA
      241 TAGACCGAGA TAGGGTTGAG TGTTGTTCCA GTTTGGAACA AGAGTCCACT ATTAAAGAAC
      301 GTGGACTCCA ACGTCAAAGG GCGAAAAACC GTCTATCAGG GCGATGGCCC ACTACGTGAA
      361 CCATCACCCT AATCAAGTTT TTTGGGGTCG AGGTGCCGTA AAGCACTAAA TCGGAACCCT
      421 AAAGGGAGCC CCCGATTTAG AGCTTGACGG GGAAAGCCGG CGAACGTGGC GAGAAAGGAA
      481 GGGAAGAAAG CGAAAGGAGC GGGCGCTAGG GCGCTGGCAA GTGTAGCGGT CACGCTGCGC
      541 GTAACCACCA CACCCGCCGC GCTTAATGCG CCGCTACAGG GCGCGTCCCA TTCGCCATTC
      601 AGGCTGCGCA ACTGTTGGGA AGGGCGATCG GTGCGGGCCT CTTCGCTATT ACGCCAGCTG
      661 GCGAAAGGGG GATGTGCTGC AAGGCGATTA AGTTGGGTAA CGCCAGGGTT TTCCCAGTCA
      721 CGACGTTGTA AAACGACGGC CAGTGAGCGC GCCTCGTTCA TTCACGTTTT TGAACCCGTG
      781 GAGGACGGGC AGACTCGCGG TGCAAATGTG TTTTACAGCG TGATGGAGCA GATGAAGATG
      841 CTCGACACGC TGCAGAACAC GCAGCTAGAT TAACCCTAGA AAGATAATCA TATTGTGACG
      901 TACGTTAAAG ATAATCATGC GTAAAATTGA CGCATGTGTT TTATCGGTCT GTATATCGAG
      961 GTTTATTTAT TAATTTGAAT AGATATTAAG TTTTATTATA TTTACACTTA CATACTAATA
     1021 ATAAATTCAA CAAACAATTT ATTTATGTTT ATTTATTTAT TAAAAAAAAA CAAAAACTCA
     1081 AAATTTCTTC TATAAAGTAA CAAAACTTTT ATGAGGGACA GCCCCCCCCC AAAGCCCCCA
     1141 GGGATGTAAT TACGTCCCTC CCCCGCTAGG GGGCAGCAGC GAGCCGCCCG GGGCTCCGCT
     1201 CCGGTCCGGC GCTCCCCCCG CATCCCCGAG CCGGCAGCGT GCGGGGACAG CCCGGGCACG
     1261 GGGAAGGTGG CACGGGATCG CTTTCCTCTG AACGCTTCTC GCTGCTCTTT GAGCCTGCAG
     1321 ACACCTGGGG GGATACGGGG AAAAGGCCTC CACgGCCaac tagtgaaaaa ccgcacttgt
     1381 ccggaaaccc cgggaatcta acccggctga acggatttag agtccgttcg atctacatga
     1441 tcaggtttcc gaggtaccca ggcggcgcac aagctatata aacctgaagg aaatctcaac
     1501 tttacactta ggtcaagtta cttatcgtac tagagcttca gcaggaaatt taactaaaat
     1561 ctaatttaac cagcatagca aatatcattt attcccaaaa tgctaaagtt tgagataaac
     1621 ggacttgatt tccggctgtt ttgacactat ccagaatgcc ttgcagatgg gtggggcatg
     1681 ctaaatactg caggaaaaac cgcacttgtc cggaaacccc gggaatctaa cccggctgaa
     1741 cggatttaga gtccgttcga tctacatgat caggtttccg aggtacccag gcggcgcaca
     1801 agctatataa acctgaagga aatctcaact ttacacttag gtcaagttac ttatcgtact
     1861 agagcttcag caggaaattt aactaaaatc taatttaacc agcatagcaa atatcattta
     1921 ttcccaaaat gctaaagttt gagataaacg gacttgattt ccggctgttt tgacactatc
     1981 cagaatgcct tgcagatggg tggggcatgc taaatactgc aggaaaaacc gcacttgtcc
     2041 ggaaaccccg ggaatctaac ccggctgaac ggatttagag tccgttcgat ctacatgatc
     2101 aggtttccga ggtacccagg cggcgcacaa gctatataaa cctgaaggaa atctcaactt
     2161 tacacttagg tcaagttact tatcgtacta gagcttcagc aggaaattta actaaaatct
     2221 aatttaacca gcatagcaaa tatcatttat tcccaaaatg ctaaagtttg agataaacgg
     2281 acttgatttc cggctgtttt gacactatcc agaatgcctt gcagatgggt ggggcatgct
     2341 aaatactgca ggaaaaaccg cacttgtccg gaaaccccgg gaatctaacc cggctgaacg
     2401 gatttagagt ccgttcgatc tacatgatca ggtttccgag gtacccaggc ggcgcacaag
     2461 ctatataaac ctgaaggaaa tctcaacttt acacttaggt caagttactt atcgtactag
     2521 agcttcagca ggaaatttaa ctaaaatcta atttaaccag catagcaaat atcatttatt
     2581 cccaaaatgc taaagtttga gataaacgga cttgatttcc ggctgttttg acactatcca
     2641 gaatgccttg cagatgggtg gggcatgcta aatactgcag aactagtgct ctagaaagga
     2701 tctgcgatcg ctccggtgcc cgtcagtggg cagagcgcac atcgcccaca gtccccgaga
     2761 agttgggggg aggggtcggc aattgaacgg gtgcctagag aaggtggcgc ggggtaaact
     2821 gggaaagtga tgtcgtgtac tggctccgcc tttttcccga gggtggggga gaaccgtata
     2881 taagtgcagt agtcgccgtg aacgttcttt ttcgcaacgg gtttgccgcc agaacacagc
     2941 tgaagcttcg aggggctcgc atctctcctt cacgcgcccg ccgccctacc tgaggccgcc
     3001 atccacgccg gttgagtcgc gttctgccgc ctcccgcctg tggtgcctcc tgaactgcgt
     3061 ccgccgtcta ggtaagttta aagctcaggt cgagaccggg cctttgtccg gcgctccctt
     3121 ggagcctacc tagactcagc cggctctcca cgctttgcct gaccctgctt gctcaactct
     3181 acgtctttgt ttcgttttct gttctgcgcc gttacagatc caagctgtga ccggcgccta
     3241 ctctagagCT AGCCACCATG GACTACAAGG ACGACGACGA CAAGccgctg cagctgccgc
     3301 cgctggaacg cctgaccctg gatatggaca aaaaaccgct gaataccctg atctctgcta
     3361 ctggtctgtg gatgagtcgt accggaacca ttcataaaat caaacaccac gaggttagcc
     3421 gttcgaaaat ctatattgag atggcgtgtg gcgatcatct ggttgtgaac aatagccgct
     3481 cttctcgtac agcacgtgca ctgcgtcacc acaaatatcg taaaacctgt aaacgttgcc
     3541 gtgtgtccga tgaggatctg aacaaattcc tgacaaaagc caatgaggac caaacaagcg
     3601 tgaaagtgaa agtcgttagc gctcctaccc gtactaaaaa agcaatgccg aaatccgttg
     3661 ctcgtgcccc taaaccactg gaaaacactg aagcagcaca ggcacagccg tctggaagca
     3721 aattctctcc ggccattcct gtttctaccc aggagtccgt ttctgttcca gcaagtgtga
     3781 gcaccagcat tagcagtatt agcaccggtg ccaccgctag cgccctggtt aaaggcaata
     3841 ccaatccgat tacaagcatg tctgccccgg ttcaagcatc agctccagca ctgacaaaat
     3901 cccaaaccga tcgtctggag gttctgctga atccgaaaga cgaaatcagc ctgaattccg
     3961 gcaaaccgtt tcgtgaactg gagagcgaac tgctgtcacg tcgtaaaaaa gacctgcaac
     4021 aaatctatgc cgaagaacgt gagaactatc tggggaaact ggaacgtgaa atcacccgct
     4081 ttttcgtgga tcgtggcttt ctggagatca aatccccgat tctgattcct ctggagtata
     4141 tcgagcgtat gggcatcgac aatgataccg aactgagcaa acaaattttc cgtgtggata
     4201 aaaacttctg tctgcgccct atgctagcac caaatctggc taactatctg cgcaaactgg
     4261 accgtgccct gcctgatcct atcaaaatct tcgagatcgg cccgtgttat cgtaaagagt
     4321 ccgacggtaa agaacatctg gaggagttta ccatgctgaa cttttgccaa atgggttcag
     4381 gttgtactcg tgagaacctg gaaagcatca tcaccgattt tctgaaccac ctgggcattg
     4441 acttcaaaat tgtgggcgac agctgtatgg tgtttggcga caccctggat gtcatgcacg
     4501 gcgacctgga actgtctagt gccgttgttg gaccaattcc gctggaccgt gagtggggta
     4561 tcgacaaacc gtggatcgga gcaggattcg gtctggaacg cctgctgaaa gtgaaacacg
     4621 acttcaaaaa catcaaacgt gccgcccgtt ctgaatcgta ttataacggg atctctacga
     4681 acctgtaata aggatccgcg gccgcgcccc tctccctccc ccccccctaa cgttactggc
     4741 cgaagccgct tggaataagg ccggtgtgcg tttgtctata tgttattttc caccatattg
     4801 ccgtcttttg gcaatgtgag ggcccggaaa cctggccctg tcttcttgac gagcattcct
     4861 aggggtcttt cccctctcgc caaaggaatg caaggtctgt tgaatgtcgt gaaggaagca
     4921 gttcctctgg aagcttcttg aagacaaaca acgtctgtag cgaccctttg caggcagcgg
     4981 aaccccccac ctggcgacag gtgcctctgc ggccaaaagc cacgtgtata agatacacct
     5041 gcaaaggcgg cacaacccca gtgccacgtt gtgagttgga tagttgtgga aagagtcaaa
     5101 tggctctcct caagcgtatt caacaagggg ctgaaggatg cccagaaggt accccattgt
     5161 atgggatctg atctggggcc tcggtgcaca tgctttacat gtgtttagtc gaggttaaaa
     5221 aaacgtctag gccccccgaa ccacggggac gtggttttcc tttgaaaaac acgatgataa
     5281 tatggccaca accatggcgt ccggaatgac cgagtacaag cccacggtgc gcctcgccac
     5341 ccgcgacgac gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc
     5401 cacgcgccac accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact
     5461 cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc
     5521 ggtggcggtc tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg
     5581 cccgcgcatg gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct
     5641 cctggcgccg caccggccca aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc
     5701 cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga
     5761 gcgcgccggg gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc ccttctacga
     5821 gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg
     5881 catgacccgc aagcccggtg ccTGAGTCGA CAATCAACCT CTGGATTACA AAATTTGTGA
     5941 AAGATTGACT GGTATTCTTA ACTATGTTGC TCCTTTTACG CTATGTGGAT ACGCTGCTTT
     6001 AATGCCTTTG TATCATGTTA ACTTGTTTAT TGCAGCTTAT AATGGTTACA AATAAAGCAA
     6061 TAGCATCACA AATTTCACAA ATAAAGCATT TTTTTCACTG CATTCTAGTT GTGGTTTGTC
     6121 CAAACTCATC AATGTATCTT ATCATGTCTG GAATTGACTC AAATGATGTC AATTAGTCTA
     6181 TCAGAAGCTA TCTGGTCTCC CTTCCGGGGG ACAAGACATC CCTGTTTAAT ATTTAAACAG
     6241 CAGTGTTCCC AAACTGGGTT CTTATATCCC TTGCTCTGGT CAACCAGGTT GCAGGGTTTC
     6301 CTGTCCTCAC AGGAACGAAG TCCCTAAAGA AACAGTGGCA GCCAGGTTTA GCCCCGGAAT
     6361 TGACTGGATT CCTTTTTTAG GGCCCATTGG TATGGCTTTT TCCCCGTATC CCCCCAGGTG
     6421 TCTGCAGGCT CAAAGAGCAG CGAGAAGCGT TCAGAGGAAA GCGATCCCGT GCCACCTTCC
     6481 CCGTGCCCGG GCTGTCCCCG CACGCTGCCG GCTCGGGGAT GCGGGGGGAG CGCCGGACCG
     6541 GAGCGGAGCC CCGGGCGGCT CGCTGCTGCC CCCTAGCGGG GGAGGGACGT AATTACATCC
     6601 CTGGGGGCTT TGGGGGGGGG CTGTCCCTGA TATCTATAAC AAGAAAATAT ATATATAATA
     6661 AGTTATCACG TAAGTAGAAC ATGAAATAAC AATATAATTA TCGTATGAGT TAAATCTTAA
     6721 AAGTCACGTA AAAGATAATC ATGCGTCATT TTGACTCACG CGGTCGTTAT AGTTCAAAAT
     6781 CAGTGACACT TACCGCATTG ACAAGCACGC CTCACGGGAG CTCCAAGCGG CGACTGAGAT
     6841 GTCCTAAATG CACAGCGACG GATTCGCGCT ATTTAGAAAG AGAGAGCAAT ATTTCAAGAA
     6901 TGCATGCGTC AATTTTACGC AGACTATCTT TCTAGGGTTA ATCTAGCTGC ATCAGGATCA
     6961 TATCGTCGGG TCTTTTTTCC GGCTCAGTCA TCGCCCAAGC TGGCGCTATC TGGGCATCGG
     7021 GGAGGAAGAA GCCCGTGCCT TTTCCCGCGA GGTTGAAGCG GCATGGAAAG AGTTTGCCGA
     7081 GGATGACTGC TGCTGCATTG ACGTTGAGCG AAAACGCACG TTTACCATGA TGATTCGGGA
     7141 AGGTGTGGCC ATGCACGCCT TTAACGGTGA ACTGTTCGTT CAGGCCACCT GGGATACCAG
     7201 TTCGTCGCGG CTTTTCCGGA CACAGTTCCG GATGGTCAGC CCGAAGCGCA TCAGCAACCC
     7261 GAACAATACC GGCGACAGCC GGAACTGCCG TGCCGGTGTG CAGATTAATG ACAGCGGTGC
     7321 GGCGCTGGGA TATTACGTCA GCGAGGACGG GTATCCTGGC TGGATGCCGC AGAAATGGAC
     7381 ATGGATACCC CGTGAGTTAC CCGGCGGGCG CGCTTGGCGT AATCATGGTC ATAGCTGTTT
     7441 CCTGTGTGAA ATTGTTATCC GCTCACAATT CCACACAACA TACGAGCCGG AAGCATAAAG
     7501 TGTAAAGCCT GGGGTGCCTA ATGAGTGAGC TAACTCACAT TAATTGCGTT GCGCTCACTG
     7561 CCCGCTTTCC AGTCGGGAAA CCTGTCGTGC CAGCTGCATT AATGAATCGG CCAACGCGCG
     7621 GGGAGAGGCG GTTTGCGTAT TGGGCGCTCT TCCGCTTCCT CGCTCACTGA CTCGCTGCGC
     7681 TCGGTCGTTC GGCTGCGGCG AGCGGTATCA GCTCACTCAA AGGCGGTAAT ACGGTTATCC
     7741 ACAGAATCAG GGGATAACGC AGGAAAGAAC ATGTGAGCAA AAGGCCAGCA AAAGGCCAGG
     7801 AACCGTAAAA AGGCCGCGTT GCTGGCGTTT TTCCATAGGC TCCGCCCCCC TGACGAGCAT
     7861 CACAAAAATC GACGCTCAAG TCAGAGGTGG CGAAACCCGA CAGGACTATA AAGATACCAG
     7921 GCGTTTCCCC CTGGAAGCTC CCTCGTGCGC TCTCCTGTTC CGACCCTGCC GCTTACCGGA
     7981 TACCTGTCCG CCTTTCTCCC TTCGGGAAGC GTGGCGCTTT CTCATAGCTC ACGCTGTAGG
     8041 TATCTCAGTT CGGTGTAGGT CGTTCGCTCC AAGCTGGGCT GTGTGCACGA ACCCCCCGTT
     8101 CAGCCCGACC GCTGCGCCTT ATCCGGTAAC TATCGTCTTG AGTCCAACCC GGTAAGACAC
     8161 GACTTATCGC CACTGGCAGC AGCCACTGGT AACAGGATTA GCAGAGCGAG GTATGTAGGC
     8221 GGTGCTACAG AGTTCTTGAA GTGGTGGCCT AACTACGGCT ACACTAGAAG GACAGTATTT
     8281 GGTATCTGCG CTCTGCTGAA GCCAGTTACC TTCGGAAAAA GAGTTGGTAG CTCTTGATCC
     8341 GGCAAACAAA CCACCGCTGG TAGCGGTGGT TTTTTTGTTT GCAAGCAGCA GATTACGCGC
     8401 AGAAAAAAAG GATCTCAAGA AGATCCTTTG ATCTTTTCTA CGGGGTCTGA CGCTCAGTGG
     8461 AACGAAAACT CACGTTAAGG GATTTTGGTC ATGAGATTAT CAAAAAGGAT CTTCACCTAG
     8521 ATCCTTTTAA ATTAAAAATG AAGTTTTAAA TCAATCTAAA GTATATATGA GTAAACTTGG
     8581 TCTGACAGTT ACCAATGCTT AATCAGTGAG GCACCTATCT CAGCGATCTG TCTATTTCGT
     8641 TCATCCATAG TTGCCTGACT CCCCGTCGTG TAGATAACTA CGATACGGGA GGGCTTACCA
     8701 TCTGGCCCCA GTGCTGCAAT GATACCGCGA GACCCACGCT CACCGGCTCC AGATTTATCA
     8761 GCAATAAACC AGCCAGCCGG AAGGGCCGAG CGCAGAAGTG GTCCTGCAAC TTTATCCGCC
     8821 TCCATCCAGT CTATTAATTG TTGCCGGGAA GCTAGAGTAA GTAGTTCGCC AGTTAATAGT
     8881 TTGCGCAACG TTGTTGCCAT TGCTACAGGC ATCGTGGTGT CACGCTCGTC GTTTGGTATG
     8941 GCTTCATTCA GCTCCGGTTC CCAACGATCA AGGCGAGTTA CATGATCCCC CATGTTGTGC
     9001 AAAAAAGCGG TTAGCTCCTT CGGTCCTCCG ATCGTTGTCA GAAGTAAGTT GGCCGCAGTG
     9061 TTATCACTCA TGGTTATGGC AGCACTGCAT AATTCTCTTA CTGTCATGCC ATCCGTAAGA
     9121 TGCTTTTCTG TGACTGGTGA GTACTCAACC AAGTCATTCT GAGAATAGTG TATGCGGCGA
     9181 CCGAGTTGCT CTTGCCCGGC GTCAATACGG GATAATACCG CGCCACATAG CAGAACTTTA
     9241 AAAGTGCTCA TCATTGGAAA ACGTTCTTCG GGGCGAAAAC TCTCAAGGAT CTTACCGCTG
     9301 TTGAGATCCA GTTCGATGTA ACCCACTCGT GCACCCAACT GATCTTCAGC ATCTTTTACT
     9361 TTCACCAGCG TTTCTGGGTG AGCAAAAACA GGAAGGCAAA ATGCCGCAAA AAAGGGAATA
     9421 AGGGCGACAC GGAAATGTTG AATACTCAT
//