LOCUS       sequence-430949-        4337 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses Methanosarcina barkeri aminoacyl-tRNA synthetases; can be 
            packaged into bacmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4337)
  AUTHORS   Jewel D, Kelemen RE, Huang RL, Zhu Z, Sundaresh B, Cao X, Malley K, 
            Huang Z, Pasha M, Anthony J, van Opijnen T, Chatterjee A
  TITLE     Virus-assisted directed evolution of enhanced suppressor tRNAs in 
            mammalian cells.
  JOURNAL   Nat Methods. 2023 Jan;20(1):95-103. doi: 10.1038/s41592-022-01706-w.
            Epub 2022 Dec 22.
  PUBMED    36550276
REFERENCE   2  (bases 1 to 4337)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/s41592-022-01706-w"; journalName: "Nat Methods"; date: 
            "2023-01"; volume: "20"; issue: "1"; pages: "95-103"
COMMENT     Sequence Label: pIDTSmart-MbPylRS
FEATURES             Location/Qualifiers
     source          1..4337
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     5..22
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6..22
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        264..643
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        644..847
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     798..818
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     892..911
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        892..910
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      930..939
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(2260..2277)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2266..2490
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2638..2654)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2638..2654)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     terminator      2702..2733
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     promoter        2734..2836
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             2837..3652
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCLKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNGDRVFRLAQAQSRMNNGLVGA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2910..2929)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      3676..3703
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     rep_origin      complement(3715..4302)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3795..3814)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cccgtgtaaa acgacggcca gtttatctag tcagcttgat tctagctgat cgtggaccgg
       61 aaggtgagcc agtgagttga ttgcagtcca gttacgctgg agtctgaggc tcgtcctgaa
      121 tgatatgcga ccgccggagg gttgcgtttg agacgggcga cagatccagt cgcgctgctc
      181 tcgtcgatcc gctagggcgg ccgcaaatac ctgcaggatc cgttttgcgc tgcttcgcga
      241 tgtacgggcc agatatacgc gttgacattg attattgact agttattaat agtaatcaat
      301 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
      361 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
      421 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
      481 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
      541 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
      601 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca
      661 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
      721 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
      781 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
      841 cagagctctc tggctaacta gagaacccac tgcttactgg cttatcgaaa ttaatacgac
      901 tcactatagg gagacccaag ctggctagcg ccaccatgga taaaaaacca ttagatgttt
      961 taatatctgc gaccgggctc tggatgtcca ggactggcac gctccacaaa atcaagcacc
     1021 atgaggtctc aagaagtaaa atatacattg aaatggcgtg tggagaccat cttgttgtga
     1081 ataattccag gagttgtaga acagccagag cattcagaca tcataagtac agaaaaacct
     1141 gcaaacgatg tagggtttcg gacgaggata tcaataattt tctcacaaga tcaaccgaaa
     1201 gcaaaaacag tgtgaaagtt agggtagttt ctgctccaaa ggtcaaaaaa gctatgccga
     1261 aatcagtttc aagggctccg aagcctctgg aaaattctgt ttctgcaaag gcatcaacga
     1321 acacatccag atctgtacct tcgcctgcaa aatcaactcc aaattcgtct gttcccgcat
     1381 cggctcctgc tccttcactt acaagaagcc agcttgatag ggttgaggct ctcttaagtc
     1441 cagaggataa aatttctcta aatatggcaa agcctttcag ggaacttgag cctgaacttg
     1501 tgacaagaag aaaaaacgat tttcagcggc tctataccaa tgatagagaa gactacctcg
     1561 gtaaactcga acgtgatatt acgaaatttt tcgtagaccg gggttttctg gagataaagt
     1621 ctcctatcct tattccggcg gaatacgtgg agagaatggg tattaataat gatactgaac
     1681 tttcaaaaca gatcttccgg gtggataaaa atctctgctt gaggccaatg cttgccccga
     1741 ctctttacaa ctatctgcga aaactcgata ggattttacc aggcccaata aaaattttcg
     1801 aagtcggacc ttgttaccgg aaagagtctg acggcaaaga gcacctggaa gaatttacta
     1861 tggtgaactt ctgtcagatg ggttcgggat gtactcggga aaatcttgaa gctctcatca
     1921 aagagtttct ggactatctg gaaatcgact tcgaaatcgt aggagattcc tgtatggtct
     1981 ttggggatac tcttgatata atgcacgggg acctggagct ttcttcggca gtcgtcgggc
     2041 cagtttctct tgatagagaa tggggtattg acaaaccatg gataggtgca ggttttggtc
     2101 ttgaacgctt gctcaaggtt atgcacggct ttaaaaacat taagagggca tcaaggtccg
     2161 aatcttacta taatgggatt tcaaccaatc tgtaagaatt caacgcgtta agtcgacttt
     2221 aactcgagtc tagagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt
     2281 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     2341 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     2401 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     2461 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc ctaggggtgc
     2521 gagcggatcg agcagtgtcg atcactactg gaccgcgagc tgtgctgcga cccgtgatct
     2581 tacggcatta tacgtatgat cggtccacga tcagctagat tatctagtca gcttgatgtc
     2641 atagctgttt cctgaggctc aatactgacc atttaaatca tacctgacct ccatagcaga
     2701 aagtcaaaag cctccgaccg gaggcttttg acttgatcgg cacgtaagag gttccaactt
     2761 tcaccataat gaaataagat cactaccggg cgtatttttt gagttatcga gattttcagg
     2821 agctaaggaa gctaaaatga gccatattca acgggaaacg tcttgcttga agccgcgatt
     2881 aaattccaac atggatgctg atttatatgg gtataaatgg gctcgcgata atgtcgggca
     2941 atcaggtgcg acaatctatc gattgtatgg gaagcccgat gcgccagagt tgtttctgaa
     3001 acatggcaaa ggtagcgttg ccaatgatgt tacagatgag atggtcaggc taaactggct
     3061 gacggaattt atgcctcttc cgaccatcaa gcattttatc cgtactcctg atgatgcatg
     3121 gttactcacc actgcgatcc cagggaaaac agcattccag gtattagaag aatatcctga
     3181 ttcaggtgaa aatattgttg atgcgctggc agtgttcctg cgccggttgc attcgattcc
     3241 tgtttgtaat tgtcctttta acggcgatcg cgtatttcgt ctcgctcagg cgcaatcacg
     3301 aatgaataac ggtttggttg gtgcgagtga ttttgatgac gagcgtaatg gctggcctgt
     3361 tgaacaagtc tggaaagaaa tgcataaact cttgccattc tcaccggatt cagtcgtcac
     3421 tcatggtgat ttctcacttg ataaccttat ttttgacgag gggaaattaa taggttgtat
     3481 tgatgttgga cgagtcggaa tcgcagaccg ataccaggat cttgccatcc tatggaactg
     3541 cctcggtgag ttttctcctt cattacagaa acggcttttt caaaaatatg gtattgataa
     3601 tcctgatatg aataaattgc agtttcactt gatgctcgat gagtttttct aatgaggacc
     3661 taaatgtaat cacctggctc accttcgggt gggcctttct gcgttgctgg cgtttttcca
     3721 taggctccgc ccccctgacg agcatcacaa aaatcgatgc tcaagtcaga ggtggcgaaa
     3781 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     3841 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     3901 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     3961 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     4021 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     4081 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     4141 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttacctcgga
     4201 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     4261 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgattttc
     4321 taccgaagaa aggccca
//
