LOCUS       sequence-448434-        8209 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Phosphoserine orthogonal translation system (preferred variant).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8209)
  AUTHORS   Pirman NL, Barber KW, Aerni HR, Ma NJ, Haimovich AD, Rogulina S, 
            Isaacs FJ, Rinehart J
  TITLE     A flexible codon in genomically recoded Escherichia coli permits 
            programmable protein phosphorylation.
  JOURNAL   Nat Commun. 2015 Sep 9;6:8130. doi: 10.1038/ncomms9130.
  PUBMED    26350500
REFERENCE   2  (bases 1 to 8209)
  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"; journalName: "Nat 
            Commun."; date: "2015-09-9"; volume: "6:8130. doi"; pages: " 
            10.1038/ncomms9130"
COMMENT     Sequence Label: SepOTS-lambda
FEATURES             Location/Qualifiers
     source          1..8209
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..78
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             79..1161
                     /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
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDDSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLATIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(98..117)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1393..1398,1399..1415,1416..1422)
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1430..1446
                     /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     1435..1457
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     complement(4396..4413)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      4566..4652
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4744..4771
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     CDS             4967..5158
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(5152..5174)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    5260..5399
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     5315..5334
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5414..5431)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5585..6173)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5665..5684)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(6284..6303)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             6295..7110
                     /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="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVAHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMSKLQFHLMLDEFF"
     primer_bind     complement(6368..6387)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 gacaccatcg aatggtgcaa aacctttcgc ggtatggcat gatagcgccc ggaagagagt
       61 caattcaggg tggtgaatgt gaaaccagta acgttatacg atgtcgcaga gtatgccggt
      121 gtctcttatc agaccgtttc ccgcgtggtg aaccaggcca gccacgtttc tgcgaaaacg
      181 cgggaaaaag tggaagcggc gatggcggag ctgaattaca ttcccaaccg cgtggcacaa
      241 caactggcgg gcaaacagtc gttgctgatt ggcgttgcca cctccagtct ggccctgcac
      301 gcgccgtcgc aaattgtcgc ggcgattaaa tctcgcgccg atcaactggg tgccagcgtg
      361 gtggtgtcga tggtagaacg aagcggcgtc gaagcctgta aagcggcggt gcacaatctt
      421 ctcgcgcaac gcgtcagtgg gctgatcatt aactatccgc tggatgacca ggatgccatt
      481 gctgtggaag ctgcctgcac taatgttccg gcgttatttc ttgatgactc tgaccagaca
      541 cccatcaaca gtattatttt ctcccatgaa gacggtacgc gactgggcgt ggagcatctg
      601 gtcgcattgg gtcaccagca aatcgcgctg ttagcgggcc cattaagttc tgtctcggcg
      661 cgtctgcgtc tggctggctg gcataaatat ctcactcgca atcaaattca gccgatagcg
      721 gaacgggaag gcgactggag tgccatgtcc ggttttcaac aaaccatgca aatgctgaat
      781 gagggcatcg ttcccactgc gatgctggtt gccaacgatc agatggcgct gggcgcaatg
      841 cgcgccatta ccgagtccgg gctgcgcgtt ggtgcggata tctcggtagt gggatacgac
      901 gataccgaag acagctcatg ttatatcccg ccgttagcca ccatcaaaca ggattttcgc
      961 ctgctggggc aaaccagcgt ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag
     1021 ggcaatcagc tgttgcccgt ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg
     1081 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     1141 cgactggaaa gcgggcagtg agcgcaacgc aattaatgta agttagcgcg aattgatctg
     1201 gtttgacagc ttatcatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc
     1261 ggaagctgtg gtatggctgt gcaggtcgta gatcactgca taattcgtgt cgctcaaggc
     1321 gcactcccgt tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc
     1381 tgaaatgagc tgttgacaat taatcatccg gcccgtataa tgtgtggaat tgtgagcgga
     1441 taacaatttc acacaggaaa cagaccatgg caatgtttaa aagagaagaa atcattgaaa
     1501 tggccaataa ggactttgaa aaagcatgga tcgaaactaa agaccttata aaagctaaaa
     1561 agataaacga aagttaccca agaataaaac cagtttttgg aaaaacacac cctgtaaatg
     1621 acactattga aaatttaaga caggcatatc ttagaatggg ttttgaagaa tatataaacc
     1681 cagtaattgt cgatgaaaga gatatttata aacaattcgg cccagaagct atggcagttt
     1741 tggatagatg cttttattta gcgggacttc caagacctga cgttggtttg agcgatgaaa
     1801 aaatttcaca gattgaaaaa cttggaatta aagtttctga gcacaaagaa agtttacaaa
     1861 aaatacttca cggatacaaa aaaggaactc ttgatggtga cgatttagtt ttagaaattt
     1921 caaatgcact tgaaatttca agcgagatgg gtttaaaaat tttagaagat gttttcccag
     1981 aatttaagga tttaaccgca gtttcttcaa aattaacttt aagaagccac atgacttcag
     2041 gatggttcct tactgtttca gacctcatga acaaaaaacc cttgccattt aaactctttt
     2101 caatcgatag atgttttaga agagaacaaa aagaagataa aagccactta atgacatacc
     2161 actctgcatc ctgtgcaatt gcaggtgaag gcgtggatat taatgatgga aaagcaattg
     2221 cagaaggatt attatcccaa tttggcttta caaactttaa attcattcct gatgaaaaga
     2281 aaagtaaata ctacacccct gaaacacaga ctgaagttta cgcataccac ccaaaattaa
     2341 aagaatggct cgaagttgct acatttggag tatattcgcc agttgcatta agcaaatacg
     2401 gaatagatgt acctgtaatg aatttgggtc ttggtgttga aagacttgca atgatttctg
     2461 gaaatttcgc agatgttcga gaaatggtat atcctcagtt ttacgaacac gaacttaatg
     2521 accgggatgt cgcttcaatg gtaaaactcg ataaagttcc agtaatggat gaaatttacg
     2581 atttaacaaa agaattaatt gagtcatgtg ttaaaaacaa agatttaaaa tccccttgtg
     2641 aattagctat tgaaaaaacg ttttcatttg gaaaaaccaa gaaaaatgta aaaataaaca
     2701 ttttttcaaa aatagaaggt aaaaatttac tcggaccttc aattttaaac gaaatctacg
     2761 tttacgatgg aaatgtaatt ggaattcctg aaagctttga cggagtaaaa gaagaattta
     2821 aagacttctt agaaaaagga aaatcagaag gggtagcaac aggcattcga tatatcgatg
     2881 cgctttgctt taaaattact tcaaaattag aagaagcatt tgtgtcaaac actactgaat
     2941 tcaaagttaa agttcgacgt gtcagaagtt taagcgacat taacttaaaa atcgatgata
     3001 tcgcaataaa acagatcatg agcaaaaata aagtaatcga cgttagaggc ccagtctttt
     3061 taaatgtcga agtaaaaatt gaataagagc tcggtaccta aagtatatta gttaagtata
     3121 agaaggagat atacatatgg gatccatgtc taaagaaaag tttgaacgta caaaaccgca
     3181 cgttaacgtc ggtactatcg gccacgttga ccatggtaaa acaacgctga ccgctgcaat
     3241 cactaccgta ctggctaaaa cctacggcgg tgctgctcgc gcattcgacc agatcgataa
     3301 cgcgccggaa gaaaaagctc gtggtatcac catcaacact tctcgggttg aatacgacac
     3361 cccgacccgt cactacgcac acgtagactg cccggggcac gccgactatg ttaaaaacat
     3421 gatcaccggt gctgcgcaga tggacggcgc gatcctggta gttgctgcga ctgacggccc
     3481 gatgccgcag actcgtgagc acatcctgct gggtcgtcag gtaggcgttc cgtacatcat
     3541 cgtgttcctg aacaaatgcg acatggttga tgacgaagag ctgctggaac tggttgaaat
     3601 ggaagttcgt gaacttctgt ctcagtacga cttcccgggc gacgacactc cgatcgttcg
     3661 tggttctgct ctgaaagcgc tggaaggcga cgcagagtgg gaagcgaaaa tcctggaact
     3721 ggctggcttc ctggattctt acattccgga accagagcgt gcgattgaca agccgttcct
     3781 gctgccgatc gtcggggtat actccatctc cggtcgtggt accgttgttt cgggtcgtgt
     3841 agaacgcggt atcatcaaag ttggtgaaga agttgaaatc gttggtatca aagagactca
     3901 gaagtctacc tgtactggcg ttgaaatgtt ccgcaaactg ctggacgaag gccgtgctgg
     3961 tgagtgggta ggtgttctgc tgcgtggtat caaacgtgaa gaaatcgaac gtggtcaggt
     4021 actggctaag ccgggcacca tcaagccgca caccaagttc gaatctgaag tgtacattct
     4081 gtccaaagat gaaggcggcc gtcatactcc gttcttcaaa ggctaccgtc cgcagttcta
     4141 cttccgtact actgacgtga ctggtaccat cgaactgccg gaaggcgtag agatggtaat
     4201 gccgggcgac aacatcaaaa tggttgttac cctgatccac ccgatcgcga tggacgacgg
     4261 tctgcgtttc gcaatccgtg aaggcggccg taccgttggc gcgggcgttg tagcaaaagt
     4321 tctgagctaa tggcgcgccg tcgacctgca ggcatgcaag cttggctgtt ttggcggatg
     4381 agagaagatt ttcagcctga tacagattaa atcagaacgc agaagcggtc tgataaaaca
     4441 gaatttgcct ggcggcagta gcgcggtggt cccacctgac cccatgccga actcagaagt
     4501 gaaaggccgt agcgccgatg gtagtgtggg gtctccccat gcgagagtag ggaactgcca
     4561 ggcatcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt
     4621 tgtcggtgaa cgctctcctg agtaggacaa atccgccggg agcggatttg aacgttgcga
     4681 agcaacggcc cggagggtgg cgggcaggac gcccgccata aactgccagg catcaaatta
     4741 agcagaaggc catcctgacg gatggccttt ttgcgtttct acaaactctt tttgtttgct
     4801 agcttttctc tggtcccgcc gcatccatac cgccagttgt ttaccctcac aacgttccag
     4861 taaccgggca tgttcatcat cagtaacccg tatcgtgagc atcctctctc gtttcatcgg
     4921 tatcattacc cccatgaaca gaaatccccc ttacacggag gcatcagtga ccaaacagga
     4981 aaaaaccgcc cttaacatgg cccgctttat cagaagccag acattaacgc ttctggagaa
     5041 actcaacgag ctggacgcgg atgaacaggc agacatctgt gaatcgcttc acgaccacgc
     5101 tgatgagctt taccgcagct gcctcgcgcg tttcggtgat gacggtgaaa acctctgaca
     5161 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc
     5221 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga cccagtcacg
     5281 tagcgatagc ggagtgtata ctggcttaac tatgcggcat cagagcagat tgtactgaga
     5341 gtgcaccatt gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac cgcatcaggc
     5401 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     5461 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     5521 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     5581 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     5641 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     5701 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     5761 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     5821 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     5881 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     5941 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     6001 ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag
     6061 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     6121 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     6181 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     6241 tggtcatgaa caataaaact gtctgcttac ataaacagta atacaagggg tgttatgagc
     6301 catattcaac gggaaacgtc ttgctctagg ccgcgattaa attccaacat ggatgctgat
     6361 ttatatgggt ataaatgggc tcgcgataat gtcgggcaat caggtgcgac aatctatcga
     6421 ttgtatggga agcccgatgc gccagagttg tttctgaaac atggcaaagg tagcgttgcc
     6481 aatgatgtta cagatgagat ggtcagacta aactggctga cggaatttat gcctcttccg
     6541 accatcaagc attttatccg tactcctgat gatgcatggt tactcaccac tgcgatcccc
     6601 gggaaaacag cattccaggt attagaagaa tatcctgatt caggtgaaaa tattgttgat
     6661 gcgctggcag tgttcctgcg ccggttgcat tcgattcctg tttgtaattg tccttttaac
     6721 agcgatcgcg tatttcgtct cgctcaggcg caatcacgaa tgaataacgg tttggttgat
     6781 gcgagtgatt ttgatgacga gcgtaatggc tggcctgttg aacaagtctg gaaagaaatg
     6841 cataaacttt tgccattctc accggattca gtcgtcgctc atggtgattt ctcacttgat
     6901 aaccttattt ttgacgaggg gaaattaata ggttgtattg atgttggacg agtcggaatc
     6961 gcagaccgat accaggatct tgccatccta tggaactgcc tcggtgagtt ttctccttca
     7021 ttacagaaac ggctttttca aaaatatggt attgataatc ctgatatgag taaattgcag
     7081 tttcatttga tgctcgatga gtttttctaa gaattaattc atgagcggat acatatttga
     7141 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccagg
     7201 ccgcgcttct ttgagcgaac gatcaaaaat aagtggcgcc ccatcaaaaa aatattctca
     7261 acataaaaaa ctttgtgtaa tacttgtaac gctgaattcg ccggggtagt ctaggggtta
     7321 ggcagcggac tctaaatccg ccttacgtgg gttcaaatcc cacccccggc tccaggatcc
     7381 tctagagtcg acctgcagat ccttagcgaa agctaaggat tttttttgcc gcggcgcttc
     7441 tttgagcgaa cgatcaaaaa taagtggcgc cccatcaaaa aaatattctc aacataaaaa
     7501 actttgtgta atacttgtaa cgctgaattc gccggggtag tctaggggtt aggcagcgga
     7561 ctctaaatcc gccttacgtg ggttcaaatc ccacccccgg ctccaggatc ctctagagtc
     7621 gacctgcaga tccttagcga aagctaagga ttttttttgc ggcccggccg cgcttctttg
     7681 agcgaacgat caaaaataag tggcgcccca tcaaaaaaat attctcaaca taaaaaactt
     7741 tgtgtaatac ttgtaacgct gaattcgccg gggtagtcta ggggttaggc agcggactct
     7801 aaatccgcct tacgtgggtt caaatcccac ccccggctcc aggatcctct agagtcgacc
     7861 tgcagatcct tagcgaaagc taaggatttt ttttgccgcg gcgcttcttt gagcgaacga
     7921 tcaaaaataa gtggcgcccc atcaaaaaaa tattctcaac ataaaaaact ttgtgtaata
     7981 cttgtaacgc tgaattcgcc ggggtagtct aggggttagg cagcggactc taaatccgcc
     8041 ttacgtgggt tcaaatccca cccccggctc caggatcctc tagagtcgac ctgcagatcc
     8101 ttagcgaaag ctaaggattt tttttgcggc cgctttcacc gtcatcaccg aaacgcgcga
     8161 ggcagcagat caattcgcgc gcgaaggcga agcggcatgc atttacgtt
//
