LOCUS       sequence-175164-        6709 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli expression of K. stuttgartiensis protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6709)
  AUTHORS   Javidpour P, Deutsch S, Mutalik VK, Hillson NJ, Petzold CJ, Keasling
            JD, Beller HR
  TITLE     Investigation of Proposed Ladderane Biosynthetic Genes from Anammox 
            Bacteria by Heterologous Expression in E. coli.
  JOURNAL   PLoS One. 2016 Mar 14;11(3):e0151087. doi: 
            10.1371/journal.pone.0151087. eCollection 2016.
  PUBMED    26975050
REFERENCE   2  (bases 1 to 6709)
  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: "PLoS One.";
            date: "2016-03-14"; volume: "11(3):e0151087. doi"; pages: " 
            10.1371/journal.pone.0151087. eCollection 2016"
COMMENT     Sequence Label: JPUB_006828
FEATURES             Location/Qualifiers
     source          1..6709
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     762..781
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(801..878)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1084..1103
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1187..1206
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1187..1205
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1206..1230
                     /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)."
     RBS             1245..1267
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2760..2777
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2830..2848)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2844..2891
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2928..3383
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3015..3034)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3225..3246
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3475..4290)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(3475..4290)
                     /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
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     4198..4217
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4282..4301
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4412..5000
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4901..4920
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5154..5171
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5186..5325
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5251..5270)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5411..5433
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5427..5618)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6427..6709,1..800))
                     /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"
ORIGIN
        1 tccgcaccaa cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga
       61 tcgttggcaa ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt
      121 tgaaaaccgg acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg
      181 cgagtgagat atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg
      241 cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc
      301 gtaccgtctt catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga
      361 aataacgccg gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc
      421 ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta
      481 caggcttcga cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg
      541 gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg
      601 gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg
      661 taattcagct ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg
      721 gcctggttca ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg
      781 tataacgtta ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat
      841 gccataccgc gaaaggtttt gcgccattcg atggtgtccg ggatctcgac gctctccctt
      901 atgcgactcc tgcattagga agcagcccag tagtaggttg aggccgttga gcaccgccgc
      961 cgcaaggaat ggtgcatgca aggagatggc gcccaacagt cccccggcca cggggcctgc
     1021 caccataccc acgccgaaac aagcgctcat gagcccgaag tggcgagccc gatcttcccc
     1081 atcggtgatg tcggcgatat aggcgccagc aaccgcacct gtggcgccgg tgatgccggc
     1141 cacgatgcgt ccggcgtaga ggatcgagat ctcgatcccg cgaaattaat acgactcact
     1201 ataggggaat tgtgagcgga taacaattcc cctctagaaa taattttgtt taactttaag
     1261 aaggagatat acatatgaag aaactggttc tgatcaaccc acatccaatg ggcaacgtgg
     1321 gcgaagaaaa tgtgagtgtt ctgaaccaaa tgccggtgaa cctgggctac ctgaaagttc
     1381 tgacgccagg taattggcaa gtggacatta tcgatgagac ccaagaatac gccattgatg
     1441 aaaatacggg cgacattacc tttggtggtg cggatctggt gggtattacc agcgtgagtt
     1501 accaagccga tcgcgcctac aagattgcga cggcgtgtcg taaacgtggt attccggtta
     1561 ttatgggtgg catccatgcc acgagtaacc cagacgaggc cgcgaaatac gttgatagcg
     1621 tggtgattcg tgaaggcatc accctgtgga gtaccatcat tgacgatttt gaaaagggtc
     1681 gcctgcgtaa gcgttatgat ggcggtctga ccccaatgga aatttactgt aatctgagcc
     1741 cagatcgcaa gtttctgaaa gataaataca aataccgtta tagcggtatt attacgaccg
     1801 cgggttgtcc atttgcctgt gaattttgta gtgtgccaca atttcagggt caaaaatatc
     1861 gcgagcgtcc agtggatgat attctgaatg aatttgaaag cattaaaggt caatatcgtg
     1921 gcctgattct gaccgacgaa aacttttacg gtcatagcaa aaagagtaat gaacgtgtgc
     1981 gtaccctgtt caaaggcatg gtggaacgcg gtatctatca gaattggttt ggtttcacca
     2041 gtctgaatat ctacaaagat gacgaaaccc tggaatacat ggcgaaaagt ggttgtgtgg
     2101 gtgtgctgat tggtattgag agcattgagg aagaaagcct gaagagtatg aacaaaaacg
     2161 tgaatctgaa aattagtgtt gaaaactact atgaggccat cgcgaacatt cgtaagcatg
     2221 gtctggccgt gtggggtacg atggtttttg gtaatgatgc cgacaccagc ggtaccttcg
     2281 aacgcgtggc cgattttatc ctggatagcc atattgatat tatgacctgc ggcatcctgt
     2341 gtccgtttat caacacccca ctgtaccacc gcctgaaccg cgagaatcgt ctgtttcgta
     2401 cgcatttccc agaagattgg aagtattata ccagtcacca cctgacctac gtgctgaaaa
     2461 agctgacgct ggatgaaatg attgaaggtt tccagtacct gtatgatcgt atctttgcca
     2521 ccgaagtgct gcgcaaacgc ttccatcgtg ccaaagacga actgaataac aacatgaatg
     2581 cggccatgtt tgcgttccgt gttaatctgg attggcagaa tgtgtaccaa catctgattc
     2641 aaaacctgaa ggatctgcat acgagtggcg actatgaagc ggccctgaag tgttatgtgg
     2701 ataccgacaa acaaggtaag ggtaaaaaac tggcgagtat cgaagccgcc ccactcgagc
     2761 accaccacca ccaccactga gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg
     2821 ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa cgggtcttga
     2881 ggggtttttt gctgaaagga ggaactatat ccggattggc gaatgggacg cgccctgtag
     2941 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     3001 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     3061 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     3121 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     3181 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     3241 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     3301 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     3361 caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg cggaacccct
     3421 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgaatta attcttagaa
     3481 aaactcatcg agcatcaaat gaaactgcaa tttattcata tcaggattat caataccata
     3541 tttttgaaaa agccgtttct gtaatgaagg agaaaactca ccgaggcagt tccataggat
     3601 ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca acatcaatac aacctattaa
     3661 tttcccctcg tcaaaaataa ggttatcaag tgagaaatca ccatgagtga cgactgaatc
     3721 cggtgagaat ggcaaaagtt tatgcatttc tttccagact tgttcaacag gccagccatt
     3781 acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg attgcgcctg
     3841 agcgagacga aatacgcgat cgctgttaaa aggacaatta caaacaggaa tcgaatgcaa
     3901 ccggcgcagg aacactgcca gcgcatcaac aatattttca cctgaatcag gatattcttc
     3961 taatacctgg aatgctgttt tcccggggat cgcagtggtg agtaaccatg catcatcagg
     4021 agtacggata aaatgcttga tggtcggaag aggcataaat tccgtcagcc agtttagtct
     4081 gaccatctca tctgtaacat cattggcaac gctacctttg ccatgtttca gaaacaactc
     4141 tggcgcatcg ggcttcccat acaatcgata gattgtcgca cctgattgcc cgacattatc
     4201 gcgagcccat ttatacccat ataaatcagc atccatgttg gaatttaatc gcggcctaga
     4261 gcaagacgtt tcccgttgaa tatggctcat aacacccctt gtattactgt ttatgtaagc
     4321 agacagtttt attgttcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     4381 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     4441 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     4501 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     4561 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     4621 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     4681 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     4741 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     4801 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     4861 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     4921 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     4981 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     5041 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     5101 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     5161 cagtgagcga ggaagcggaa gagcgcctga tgcggtattt tctccttacg catctgtgcg
     5221 gtatttcaca ccgcaatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     5281 ccagtataca ctccgctatc gctacgtgac tgggtcatgg ctgcgccccg acacccgcca
     5341 acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct
     5401 gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg
     5461 aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg attcacagat gtctgcctgt
     5521 tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta atgtctggct tctgataaag
     5581 cgggccatgt taagggcggt tttttcctgt ttggtcactg atgcctccgt gtaaggggga
     5641 tttctgttca tgggggtaat gataccgatg aaacgagaga ggatgctcac gatacgggtt
     5701 actgatgatg aacatgcccg gttactggaa cgttgtgagg gtaaacaact ggcggtatgg
     5761 atgcggcggg accagagaaa aatcactcag ggtcaatgcc agcgcttcgt taatacagat
     5821 gtaggtgttc cacagggtag ccagcagcat cctgcgatgc agatccggaa cataatggtg
     5881 cagggcgctg acttccgcgt ttccagactt tacgaaacac ggaaaccgaa gaccattcat
     5941 gttgttgctc aggtcgcaga cgttttgcag cagcagtcgc ttcacgttcg ctcgcgtatc
     6001 ggtgattcat tctgctaacc agtaaggcaa ccccgccagc ctagccgggt cctcaacgac
     6061 aggagcacga tcatgcgcac ccgtggggcc gccatgccgg cgataatggc ctgcttctcg
     6121 ccgaaacgtt tggtggcggg accagtgacg aaggcttgag cgagggcgtg caagattccg
     6181 aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc gaaagcggtc ctcgccgaaa
     6241 atgacccaga gcgctgccgg cacctgtcct acgagttgca tgataaagaa gacagtcata
     6301 agtgcggcga cgatagtcat gccccgcgcc caccggaagg agctgactgg gttgaaggct
     6361 ctcaagggca tcggtcgaga tcccggtgcc taatgagtga gctaacttac attaattgcg
     6421 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     6481 ggccaacgcg cggggagagg cggtttgcgt attgggcgcc agggtggttt ttcttttcac
     6541 cagtgagacg ggcaacagct gattgccctt caccgcctgg ccctgagaga gttgcagcaa
     6601 gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt ttgatggtgg ttaacggcgg
     6661 gatataacat gagctgtctt cggtatcgtc gtatcccact accgagata
//
