LOCUS       sequence-368321-        6418 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pET-28a vector (Novagen) (encoding an N-terminal His tag), to 
            transform Escherichia coli BL21 (DE3) cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6418)
  AUTHORS   Aripirala S, Gonzalez-Pacanowska D, Oldfield E, Kaiser M, Amzel LM, 
            Gabelli SB
  TITLE     Structural and thermodynamic basis of the inhibition of Leishmania 
            major farnesyl diphosphate synthase by nitrogen-containing 
            bisphosphonates.
  JOURNAL   Acta Crystallogr D Biol Crystallogr. 2014 Mar;70(Pt 3):802-10. doi: 
            10.1107/S1399004713033221. Epub 2014 Feb 22.
  PUBMED    24598749
REFERENCE   2  (bases 1 to 6418)
  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: "Acta 
            Crystallogr D Biol Crystallogr."; date: "2014-03"; volume: "70(Pt 
            3):802-10. doi"; pages: " 10.1107/S1399004713033221. Epub 2014 Feb 
            22"
COMMENT     Sequence Label: pET28_LmFPPS
FEATURES             Location/Qualifiers
     source          1..6418
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1296..1313)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(1323..1340)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             1359..1381
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1396..1420
                     /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     complement(1420..1439)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1421..1439)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1523..1542)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1748..1825
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1826..2908
                     /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"
     primer_bind     complement(1845..1864)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3717..3908
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3902..3924)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4010..4149
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4065..4084
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4164..4181)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4335..4923)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4415..4434)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(5034..5053)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             5045..5860
                     /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             5045..5860
                     /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     complement(5118..5137)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(5952..6407)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6089..6110)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6301..6320
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagcttgt
      181 cgacggagct cgaattcgga tccttacttt tggcgcttgt aagtcttgct ccaaagcacc
      241 ttcacgctag ccgcgaaggc ggcattctgc gcctccagcg cagcgatgag ctgctcgacc
      301 tccgccacaa cggccttctc gtactcctcg aagcgggcga gcaggttctg ctcggtgtaa
      361 agctgcttga taacggcgac cgcggccgga tcagtggagc cgtagttggc cttgaactcc
      421 gcgaccttct cggccggggc ggtcgtcagg aaggtgacag cgagccacga gcacttggcg
      481 tcttcaatgt cggtgccgat tttgccaagc ttctcgggcg gggtgaagca gtccatcacg
      541 tcgtcctgca cctgaaagta ctcgcccatc accatggcca ccttgtgcgt cgccttcttg
      601 tccactttct ccagtgtccc gctcaccaga aggcccatca ccagcggcag ccagtaggtg
      661 tagtaggcgg tcttgtacac aacgatgcgg cggtggttaa agggtgtgta ctcgacgtag
      721 tccgtcgtgt tggcgtgcgc cactttggcg tccagcttcg ctgagtccac catcgacgtg
      781 acgtcgtaca gctgaccaat cgtggtagtg agatcgacgt cgtggaaaac acgcagcacc
      841 tcggcgagga acgggcggtc ggcgaagtag tgcagtgcca tctgcgtcgc ccaggccagc
      901 agaatgagac cgtcattgat ggccacctgc gcggtcacgc cggggtgcag gtaccagcac
      961 ggcttgccgc ggcgagtctt gctgtggtcc atgatgtcgt cctccacaag gaagtgggcc
     1021 tgcagcatct cgatcatcca gccgcacaca cacgcgtcgt gcagaacgcg ctccatcgca
     1081 gccgcatcca tctgcgtatc cttcgccatc gcctccgcca cgtccacgac gcagagacct
     1141 cggttgtact tgccgccgag gcacgtagtg tccatcatag acttcaggta gcccttgcgg
     1201 tggacatcca tttcgaagcg cttctccgca tcgccgagta ggaactcctg aacctcctcg
     1261 tacaccttct ggaagcgttc catgtgcgcc atatggctgc cgcgcggcac caggccgctg
     1321 ctgtgatgat gatgatgatg gctgctgccc atggtatatc tccttcttaa agttaaacaa
     1381 aattatttct agaggggaat tgttatccgc tcacaattcc cctatagtga gtcgtattaa
     1441 tttcgcggga tcgagatctc gatcctctac gccggacgca tcgtggccgg catcaccggc
     1501 gccacaggtg cggttgctgg cgcctatatc gccgacatca ccgatgggga agatcgggct
     1561 cgccacttcg ggctcatgag cgcttgtttc ggcgtgggta tggtggcagg ccccgtggcc
     1621 gggggactgt tgggcgccat ctccttgcat gcaccattcc ttgcggcggc ggtgctcaac
     1681 ggcctcaacc tactactggg ctgcttccta atgcaggagt cgcataaggg agagcgtcga
     1741 gatcccggac accatcgaat ggcgcaaaac ctttcgcggt atggcatgat agcgcccgga
     1801 agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg tcgcagagta
     1861 tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac caggccagcc acgtttctgc
     1921 gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg aattacattc ccaaccgcgt
     1981 ggcacaacaa ctggcgggca aacagtcgtt gctgattggc gttgccacct ccagtctggc
     2041 cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc aactgggtgc
     2101 cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag cggcggtgca
     2161 caatcttctc gcgcaacgcg tcagtgggct gatcattaac tatccgctgg atgaccagga
     2221 tgccattgct gtggaagctg cctgcactaa tgttccggcg ttatttcttg atgtctctga
     2281 ccagacaccc atcaacagta ttattttctc ccatgaagac ggtacgcgac tgggcgtgga
     2341 gcatctggtc gcattgggtc accagcaaat cgcgctgtta gcgggcccat taagttctgt
     2401 ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcgcaatc aaattcagcc
     2461 gatagcggaa cgggaaggcg actggagtgc catgtccggt tttcaacaaa ccatgcaaat
     2521 gctgaatgag ggcatcgttc ccactgcgat gctggttgcc aacgatcaga tggcgctggg
     2581 cgcaatgcgc gccattaccg agtccgggct gcgcgttggt gcggatatct cggtagtggg
     2641 atacgacgat accgaagaca gctcatgtta tatcccgccg ttaaccacca tcaaacagga
     2701 ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg caactctctc agggccaggc
     2761 ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa agaaaaacca ccctggcgcc
     2821 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     2881 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtaagt tagctcactc
     2941 attaggcacc gggatctcga ccgatgccct tgagagcctt caacccagtc agctccttcc
     3001 ggtgggcgcg gggcatgact atcgtcgccg cacttatgac tgtcttcttt atcatgcaac
     3061 tcgtaggaca ggtgccggca gcgctctggg tcattttcgg cgaggaccgc tttcgctgga
     3121 gcgcgacgat gatcggcctg tcgcttgcgg tattcggaat cttgcacgcc ctcgctcaag
     3181 ccttcgtcac tggtcccgcc accaaacgtt tcggcgagaa gcaggccatt atcgccggca
     3241 tggcggcccc acgggtgcgc atgatcgtgc tcctgtcgtt gaggacccgg ctaggctggc
     3301 ggggttgcct tactggttag cagaatgaat caccgatacg cgagcgaacg tgaagcgact
     3361 gctgctgcaa aacgtctgcg acctgagcaa caacatgaat ggtcttcggt ttccgtgttt
     3421 cgtaaagtct ggaaacgcgg aagtcagcgc cctgcaccat tatgttccgg atctgcatcg
     3481 caggatgctg ctggctaccc tgtggaacac ctacatctgt attaacgaag cgctggcatt
     3541 gaccctgagt gatttttctc tggtcccgcc gcatccatac cgccagttgt ttaccctcac
     3601 aacgttccag taaccgggca tgttcatcat cagtaacccg tatcgtgagc atcctctctc
     3661 gtttcatcgg tatcattacc cccatgaaca gaaatccccc ttacacggag gcatcagtga
     3721 ccaaacagga aaaaaccgcc cttaacatgg cccgctttat cagaagccag acattaacgc
     3781 ttctggagaa actcaacgag ctggacgcgg atgaacaggc agacatctgt gaatcgcttc
     3841 acgaccacgc tgatgagctt taccgcagct gcctcgcgcg tttcggtgat gacggtgaaa
     3901 acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga
     3961 gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc gcagccatga
     4021 cccagtcacg tagcgatagc ggagtgtata ctggcttaac tatgcggcat cagagcagat
     4081 tgtactgaga gtgcaccatt gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
     4141 cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     4201 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     4261 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     4321 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     4381 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     4441 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     4501 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     4561 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     4621 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     4681 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     4741 ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg
     4801 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     4861 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     4921 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     4981 taagggattt tggtcatgaa caataaaact gtctgcttac ataaacagta atacaagggg
     5041 tgttatgagc catattcaac gggaaacgtc ttgctctagg ccgcgattaa attccaacat
     5101 ggatgctgat ttatatgggt ataaatgggc tcgcgataat gtcgggcaat caggtgcgac
     5161 aatctatcga ttgtatggga agcccgatgc gccagagttg tttctgaaac atggcaaagg
     5221 tagcgttgcc aatgatgtta cagatgagat ggtcagacta aactggctga cggaatttat
     5281 gcctcttccg accatcaagc attttatccg tactcctgat gatgcatggt tactcaccac
     5341 tgcgatcccc gggaaaacag cattccaggt attagaagaa tatcctgatt caggtgaaaa
     5401 tattgttgat gcgctggcag tgttcctgcg ccggttgcat tcgattcctg tttgtaattg
     5461 tccttttaac agcgatcgcg tatttcgtct cgctcaggcg caatcacgaa tgaataacgg
     5521 tttggttgat gcgagtgatt ttgatgacga gcgtaatggc tggcctgttg aacaagtctg
     5581 gaaagaaatg cataaacttt tgccattctc accggattca gtcgtcactc atggtgattt
     5641 ctcacttgat aaccttattt ttgacgaggg gaaattaata ggttgtattg atgttggacg
     5701 agtcggaatc gcagaccgat accaggatct tgccatccta tggaactgcc tcggtgagtt
     5761 ttctccttca ttacagaaac ggctttttca aaaatatggt attgataatc ctgatatgaa
     5821 taaattgcag tttcatttga tgctcgatga gtttttctaa gaattaattc atgagcggat
     5881 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     5941 aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     6001 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     6061 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     6121 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     6181 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     6241 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     6301 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     6361 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc attcgcca
//
