LOCUS       sequence-271465-        6876 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 6876)
  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 6876)
  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..6876
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             100..117
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             127..144
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             148..180
                     /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             join(190..192,193..903)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(190..192,193..903)
                     /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(232..253)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(493..512)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     840..861
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1741..1758
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1811..1829)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1825..1872
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1909..2364
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1996..2015)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2206..2227
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2456..3271)
                     /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(2456..3271)
                     /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     3179..3198
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3263..3282
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3393..3981
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3882..3901
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4135..4152
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4167..4306
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4232..4251)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4392..4414
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4408..4599)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5408..6490)
                     /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     6452..6471
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6491..6568)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6774..6793
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atataccatg ggcagcagcc atcatcatca tcatcacagc
      121 agcggcctgg tgccgcgcgg cagccatatg gctagcatga ctggtggaca gcaaatgggt
      181 cgcggatccg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag
      241 ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc
      301 acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg
      361 cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac
      421 atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc
      481 atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac
      541 accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg
      601 gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc cgacaagcag
      661 aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag
      721 ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac
      781 aaccactacc tgagcaccca gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac
      841 atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac
      901 aaggaattcg cacgcattat tgttgttact tcgggcaaag ggggtgttgg taagacaacc
      961 tccagcgcgg ccatcgccac tggtttggcc cagaagggaa agaaaactgt cgtgatagat
     1021 tttgatatcg gcctgcgtaa tctcgacctg attatgggtt gtgaacgccg ggtcgtttac
     1081 gatttcgtca acgtcattca gggcgatgca acgctaaatc aggcgttaat taaagataag
     1141 cgtactgaaa atctctatat tctgccggca tcgcaaacac gcgataaaga tgccctcacc
     1201 cgtgaagggg tcgccaaagt tcttgatgat ctgaaagcga tggattttga atttatcgtt
     1261 tgtgactccc cggcagggat tgaaaccggt gcgttaatgg cactctattt tgcagacgaa
     1321 gccattatta ccaccaaccc ggaagtctcc tcagtacgcg actctgaccg tattttaggc
     1381 attctggcgt cgaaatcacg ccgcgcagaa aatggcgaag agcctattaa agagcacctg
     1441 ctgttaacgc gctataaccc aggccgcgta agcagaggtg acatgctgag catggaagat
     1501 gtgctggaga tcctgcgcat caaactcgtc ggcgtgatcc cagaggatca atcagtattg
     1561 cgcgcctcta accagggtga accggtcatt ctcgacatta acgccgatgc gggtaaagcc
     1621 tacgcagata ccgtagaacg tctgttggga gaagaacgtc ctttccgctt cattgaagaa
     1681 gagaagaaag gcttcctcaa acgcttgttc ggaggataaa agcttgcggc cgcactcgag
     1741 caccaccacc accaccactg agatccggct gctaacaaag cccgaaagga agctgagttg
     1801 gctgctgcca ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg
     1861 aggggttttt tgctgaaagg aggaactata tccggattgg cgaatgggac gcgccctgta
     1921 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     1981 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     2041 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     2101 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     2161 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     2221 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     2281 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     2341 acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     2401 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgaatt aattcttaga
     2461 aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta tcaataccat
     2521 atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag ttccatagga
     2581 tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata caacctatta
     2641 atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg acgactgaat
     2701 ccggtgagaa tggcaaaagt ttatgcattt ctttccagac ttgttcaaca ggccagccat
     2761 tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt gattgcgcct
     2821 gagcgagacg aaatacgcga tcgctgttaa aaggacaatt acaaacagga atcgaatgca
     2881 accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca ggatattctt
     2941 ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat gcatcatcag
     3001 gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc cagtttagtc
     3061 tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc agaaacaact
     3121 ctggcgcatc gggcttccca tacaatcgat agattgtcgc acctgattgc ccgacattat
     3181 cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat cgcggcctag
     3241 agcaagacgt ttcccgttga atatggctca taacacccct tgtattactg tttatgtaag
     3301 cagacagttt tattgttcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     3361 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     3421 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     3481 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc
     3541 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     3601 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     3661 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     3721 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     3781 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     3841 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     3901 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     3961 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     4021 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     4081 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag
     4141 tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc
     4201 ggtatttcac accgcaatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa
     4261 gccagtatac actccgctat cgctacgtga ctgggtcatg gctgcgcccc gacacccgcc
     4321 aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc
     4381 tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc
     4441 gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg
     4501 ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt aatgtctggc ttctgataaa
     4561 gcgggccatg ttaagggcgg ttttttcctg tttggtcact gatgcctccg tgtaaggggg
     4621 atttctgttc atgggggtaa tgataccgat gaaacgagag aggatgctca cgatacgggt
     4681 tactgatgat gaacatgccc ggttactgga acgttgtgag ggtaaacaac tggcggtatg
     4741 gatgcggcgg gaccagagaa aaatcactca gggtcaatgc cagcgcttcg ttaatacaga
     4801 tgtaggtgtt ccacagggta gccagcagca tcctgcgatg cagatccgga acataatggt
     4861 gcagggcgct gacttccgcg tttccagact ttacgaaaca cggaaaccga agaccattca
     4921 tgttgttgct caggtcgcag acgttttgca gcagcagtcg cttcacgttc gctcgcgtat
     4981 cggtgattca ttctgctaac cagtaaggca accccgccag cctagccggg tcctcaacga
     5041 caggagcacg atcatgcgca cccgtggggc cgccatgccg gcgataatgg cctgcttctc
     5101 gccgaaacgt ttggtggcgg gaccagtgac gaaggcttga gcgagggcgt gcaagattcc
     5161 gaataccgca agcgacaggc cgatcatcgt cgcgctccag cgaaagcggt cctcgccgaa
     5221 aatgacccag agcgctgccg gcacctgtcc tacgagttgc atgataaaga agacagtcat
     5281 aagtgcggcg acgatagtca tgccccgcgc ccaccggaag gagctgactg ggttgaaggc
     5341 tctcaagggc atcggtcgag atcccggtgc ctaatgagtg agctaactta cattaattgc
     5401 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     5461 cggccaacgc gcggggagag gcggtttgcg tattgggcgc cagggtggtt tttcttttca
     5521 ccagtgagac gggcaacagc tgattgccct tcaccgcctg gccctgagag agttgcagca
     5581 agcggtccac gctggtttgc cccagcaggc gaaaatcctg tttgatggtg gttaacggcg
     5641 ggatataaca tgagctgtct tcggtatcgt cgtatcccac taccgagata tccgcaccaa
     5701 cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa
     5761 ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg
     5821 acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat
     5881 atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca
     5941 gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt
     6001 catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg
     6061 gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa
     6121 tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga
     6181 cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt
     6241 taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa
     6301 tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct
     6361 ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca
     6421 ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta
     6481 ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc
     6541 gaaaggtttt gcgccattcg atggtgtccg ggatctcgac gctctccctt atgcgactcc
     6601 tgcattagga agcagcccag tagtaggttg aggccgttga gcaccgccgc cgcaaggaat
     6661 ggtgcatgca aggagatggc gcccaacagt cccccggcca cggggcctgc caccataccc
     6721 acgccgaaac aagcgctcat gagcccgaag tggcgagccc gatcttcccc atcggtgatg
     6781 tcggcgatat aggcgccagc aaccgcacct gtggcgccgg tgatgccggc cacgatgcgt
     6841 ccggcgtaga ggatcgagat ctcgatcccg cgaaat
//
