LOCUS       sequence-257348-        6880 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression vector for the Escherichia coli MinD with an N-terminal 
            fusion of a His-tag for purification and an EGFP for visualization..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6880)
  AUTHORS   Zieske K, Schweizer J, Schwille P
  TITLE     Surface topology assisted alignment of Min protein waves.
  JOURNAL   FEBS Lett. 2014 Aug 1;588(15):2545-9. doi: 
            10.1016/j.febslet.2014.06.026. Epub 2014 Jun 14.
  PUBMED    24937143
REFERENCE   2  (bases 1 to 6880)
  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"; doi: 
            "10.1016/j.febslet.2014.06.026"; journalName: "FEBS Lett"; date: 
            "2014-08-1- 1"; volume: "588"; issue: "15"; pages: "2545-9"
COMMENT     Sequence Label: pET28a-eGFP-MinD
FEATURES             Location/Qualifiers
     source          1..6880
                     /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(join(995..1705,1706..1708))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             complement(join(995..1705,1706..1708))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1037..1058)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1386..1405
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1645..1666
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             complement(1718..1750)
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             complement(1754..1771)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(1781..1798)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             1817..1839
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1854..1878
                     /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(1878..1897)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1879..1897)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1981..2000)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        2206..2283
                     /gene="lacI"
                     /label=lacI promoter
     CDS             2284..3366
                     /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(2303..2322)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4175..4366
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(4360..4382)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4468..4610
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4523..4542
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4625..4642)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4796..5384)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4876..4895)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(5495..5514)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             5506..6321
                     /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             5506..6321
                     /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(5579..5598)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(6414..6869)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6551..6572)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6763..6782
                     /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 gcaagctttt
      181 atcctccgaa caagcgtttg aggaagcctt tcttctcttc ttcaatgaag cggaaaggac
      241 gttcttctcc caacagacgt tctacggtat ctgcgtaggc tttacccgca tcggcgttaa
      301 tgtcgagaat gaccggttca ccctggttag aggcgcgcaa tactgattga tcctctggga
      361 tcacgccgac gagtttgatg cgcaggatct ccagcacatc ttccatgctc agcatgtcac
      421 ctctgcttac gcggcctggg ttatagcgcg ttaacagcag gtgctcttta ataggctctt
      481 cgccattttc tgcgcggcgt gatttcgacg ccagaatgcc taaaatacgg tcagagtcgc
      541 gtactgagga gacttccggg ttggtggtaa taatggcttc gtctgcaaaa tagagtgcca
      601 ttaacgcacc ggtttcaatc cctgccgggg agtcacaaac gataaattca aaatccatcg
      661 ctttcagatc atcaagaact ttggcgaccc cttcacgggt gagggcatct ttatcgcgtg
      721 tttgcgatgc cggcagaata tagagatttt cagtacgctt atctttaatt aacgcctgat
      781 ttagcgttgc atcgccctga atgacgttga cgaaatcgta aacgacccgg cgttcacaac
      841 ccataatcag gtcgagatta cgcaggccga tatcaaaatc tatcacgaca gttttctttc
      901 ccttctgggc caaaccagtg gcgatggccg cgctggaggt tgtcttacca acaccccctt
      961 tgcccgaagt aacaacaata atgcgtgcga attccttgta cagctcgtcc atgccgagag
     1021 tgatcccggc ggcggtcacg aactccagca ggaccatgtg atcgcgcttc tcgttggggt
     1081 ctttgctcag ggcggactgg gtgctcaggt agtggttgtc gggcagcagc acggggccgt
     1141 cgccgatggg ggtgttctgc tggtagtggt cggcgagctg cacgctgccg tcctcgatgt
     1201 tgtggcggat cttgaagttc accttgatgc cgttcttctg cttgtcggcc atgatataga
     1261 cgttgtggct gttgtagttg tactccagct tgtgccccag gatgttgccg tcctccttga
     1321 agtcgatgcc cttcagctcg atgcggttca ccagggtgtc gccctcgaac ttcacctcgg
     1381 cgcgggtctt gtagttgccg tcgtccttga agaagatggt gcgctcctgg acgtagcctt
     1441 cgggcatggc ggacttgaag aagtcgtgct gcttcatgtg gtcggggtag cggctgaagc
     1501 actgcacgcc gtaggtcagg gtggtcacga gggtgggcca gggcacgggc agcttgccgg
     1561 tggtgcagat gaacttcagg gtcagcttgc cgtaggtggc atcgccctcg ccctcgccgg
     1621 acacgctgaa cttgtggccg tttacgtcgc cgtccagctc gaccaggatg ggcaccaccc
     1681 cggtgaacag ctcctcgccc ttgctcacgg atccgcgacc catttgctgt ccaccagtca
     1741 tgctagccat atggctgccg cgcggcacca ggccgctgct gtgatgatga tgatgatggc
     1801 tgctgcccat ggtatatctc cttcttaaag ttaaacaaaa ttatttctag aggggaattg
     1861 ttatccgctc acaattcccc tatagtgagt cgtattaatt tcgcgggatc gagatctcga
     1921 tcctctacgc cggacgcatc gtggccggca tcaccggcgc cacaggtgcg gttgctggcg
     1981 cctatatcgc cgacatcacc gatggggaag atcgggctcg ccacttcggg ctcatgagcg
     2041 cttgtttcgg cgtgggtatg gtggcaggcc ccgtggccgg gggactgttg ggcgccatct
     2101 ccttgcatgc accattcctt gcggcggcgg tgctcaacgg cctcaaccta ctactgggct
     2161 gcttcctaat gcaggagtcg cataagggag agcgtcgaga tcccggacac catcgaatgg
     2221 cgcaaaacct ttcgcggtat ggcatgatag cgcccggaag agagtcaatt cagggtggtg
     2281 aatgtgaaac cagtaacgtt atacgatgtc gcagagtatg ccggtgtctc ttatcagacc
     2341 gtttcccgcg tggtgaacca ggccagccac gtttctgcga aaacgcggga aaaagtggaa
     2401 gcggcgatgg cggagctgaa ttacattccc aaccgcgtgg cacaacaact ggcgggcaaa
     2461 cagtcgttgc tgattggcgt tgccacctcc agtctggccc tgcacgcgcc gtcgcaaatt
     2521 gtcgcggcga ttaaatctcg cgccgatcaa ctgggtgcca gcgtggtggt gtcgatggta
     2581 gaacgaagcg gcgtcgaagc ctgtaaagcg gcggtgcaca atcttctcgc gcaacgcgtc
     2641 agtgggctga tcattaacta tccgctggat gaccaggatg ccattgctgt ggaagctgcc
     2701 tgcactaatg ttccggcgtt atttcttgat gtctctgacc agacacccat caacagtatt
     2761 attttctccc atgaagacgg tacgcgactg ggcgtggagc atctggtcgc attgggtcac
     2821 cagcaaatcg cgctgttagc gggcccatta agttctgtct cggcgcgtct gcgtctggct
     2881 ggctggcata aatatctcac tcgcaatcaa attcagccga tagcggaacg ggaaggcgac
     2941 tggagtgcca tgtccggttt tcaacaaacc atgcaaatgc tgaatgaggg catcgttccc
     3001 actgcgatgc tggttgccaa cgatcagatg gcgctgggcg caatgcgcgc cattaccgag
     3061 tccgggctgc gcgttggtgc ggatatctcg gtagtgggat acgacgatac cgaagacagc
     3121 tcatgttata tcccgccgtt aaccaccatc aaacaggatt ttcgcctgct ggggcaaacc
     3181 agcgtggacc gcttgctgca actctctcag ggccaggcgg tgaagggcaa tcagctgttg
     3241 cccgtctcac tggtgaaaag aaaaaccacc ctggcgccca atacgcaaac cgcctctccc
     3301 cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact ggaaagcggg
     3361 cagtgagcgc aacgcaatta atgtaagtta gctcactcat taggcaccgg gatctcgacc
     3421 gatgcccttg agagccttca acccagtcag ctccttccgg tgggcgcggg gcatgactat
     3481 cgtcgccgca cttatgactg tcttctttat catgcaactc gtaggacagg tgccggcagc
     3541 gctctgggtc attttcggcg aggaccgctt tcgctggagc gcgacgatga tcggcctgtc
     3601 gcttgcggta ttcggaatct tgcacgccct cgctcaagcc ttcgtcactg gtcccgccac
     3661 caaacgtttc ggcgagaagc aggccattat cgccggcatg gcggccccac gggtgcgcat
     3721 gatcgtgctc ctgtcgttga ggacccggct aggctggcgg ggttgcctta ctggttagca
     3781 gaatgaatca ccgatacgcg agcgaacgtg aagcgactgc tgctgcaaaa cgtctgcgac
     3841 ctgagcaaca acatgaatgg tcttcggttt ccgtgtttcg taaagtctgg aaacgcggaa
     3901 gtcagcgccc tgcaccatta tgttccggat ctgcatcgca ggatgctgct ggctaccctg
     3961 tggaacacct acatctgtat taacgaagcg ctggcattga ccctgagtga tttttctctg
     4021 gtcccgccgc atccataccg ccagttgttt accctcacaa cgttccagta accgggcatg
     4081 ttcatcatca gtaacccgta tcgtgagcat cctctctcgt ttcatcggta tcattacccc
     4141 catgaacaga aatccccctt acacggaggc atcagtgacc aaacaggaaa aaaccgccct
     4201 taacatggcc cgctttatca gaagccagac attaacgctt ctggagaaac tcaacgagct
     4261 ggacgcggat gaacaggcag acatctgtga atcgcttcac gaccacgctg atgagcttta
     4321 ccgcagctgc ctcgcgcgtt tcggtgatga cggtgaaaac ctctgacaca tgcagctccc
     4381 ggagacggtc acagcttgtc tgtaagcgga tgccgggagc agacaagccc gtcagggcgc
     4441 gtcagcgggt gttggcgggt gtcggggcgc agccatgacc cagtcacgta gcgatagcgg
     4501 agtgtatact ggcttaacta tgcggcatca gagcagattg tactgagagt gcaccatata
     4561 tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg cgctcttccg
     4621 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     4681 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     4741 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     4801 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     4861 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     4921 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     4981 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     5041 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     5101 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     5161 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     5221 acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     5281 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     5341 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     5401 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     5461 acaataaaac tgtctgctta cataaacagt aatacaaggg gtgttatgag ccatattcaa
     5521 cgggaaacgt cttgctctag gccgcgatta aattccaaca tggatgctga tttatatggg
     5581 tataaatggg ctcgcgataa tgtcgggcaa tcaggtgcga caatctatcg attgtatggg
     5641 aagcccgatg cgccagagtt gtttctgaaa catggcaaag gtagcgttgc caatgatgtt
     5701 acagatgaga tggtcagact aaactggctg acggaattta tgcctcttcc gaccatcaag
     5761 cattttatcc gtactcctga tgatgcatgg ttactcacca ctgcgatccc cgggaaaaca
     5821 gcattccagg tattagaaga atatcctgat tcaggtgaaa atattgttga tgcgctggca
     5881 gtgttcctgc gccggttgca ttcgattcct gtttgtaatt gtccttttaa cagcgatcgc
     5941 gtatttcgtc tcgctcaggc gcaatcacga atgaataacg gtttggttga tgcgagtgat
     6001 tttgatgacg agcgtaatgg ctggcctgtt gaacaagtct ggaaagaaat gcataaactt
     6061 ttgccattct caccggattc agtcgtcact catggtgatt tctcacttga taaccttatt
     6121 tttgacgagg ggaaattaat aggttgtatt gatgttggac gagtcggaat cgcagaccga
     6181 taccaggatc ttgccatcct atggaactgc ctcggtgagt tttctccttc attacagaaa
     6241 cggctttttc aaaaatatgg tattgataat cctgatatga ataaattgca gtttcatttg
     6301 atgctcgatg agtttttcta agaattaatt catgagcgga tacatatttg aatgtattta
     6361 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgaaattgt
     6421 aaacgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa
     6481 ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt
     6541 gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa
     6601 agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag
     6661 ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt
     6721 tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg
     6781 agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc
     6841 cgcgcttaat gcgccgctac agggcgcgtc ccattcgcca
//
