LOCUS       sequence-448069-        5914 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Bacterial expression of Vipp1 H1-6 with C-terminal 6x His-Tag and 3C
            cleavage site.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5914)
  TITLE     Structural basis for Vipp1 membrane binding: From loose coats and 
            carpets to ring and rod assemblies
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5914)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pET50delCT_Vipp1_H1-6
FEATURES             Location/Qualifiers
     source          1..5914
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(149..168)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             160..975
                     /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             160..975
                     /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(233..252)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(1067..1522)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1204..1225)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1416..1435
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(1559..1606)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     1602..1620
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(1701..1745)
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     CDS             complement(1746..1763)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(1837..1854)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1855..1878)
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     RBS             2621..2643
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    2658..2682
                     /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(2682..2701)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2683..2701)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2789..2808)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        3014..3091
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3092..4174
                     /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
                     NYIPNRVAQQLAGKQSLLIGVTTSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(3111..3130)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             4746..4937
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(4931..4953)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    5039..5178
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     5094..5113
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5193..5210)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(5364..5914,1..38))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5444..5463)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gcaagcagca gattacgcgc agaaaaaaag gatttcaaga agatcctttg atcttttcta
       61 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgaacaata
      121 aaactgtctg cttacataaa cagtaataca aggggtgtta tgagccatat tcaacgggaa
      181 acgtcttgct ctaggccgcg attaaattcc aacatggatg ctgatttata tgggtataaa
      241 tgggctcgcg ataatgtcgg gcaatcaggt gcgacaatct atcgattgta tgggaagccc
      301 gatgcgccag agttgtttct gaaacatggc aaaggtagcg ttgccaatga tgttacagat
      361 gagatggtca gactaaactg gctgacggaa tttatgcctc ttccgaccat caagcatttt
      421 atccgtactc ctgatgatgc atggttactc accactgcga tccccggcaa aacagcattc
      481 caggtattag aagaatatcc tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc
      541 ctgcgccggt tgcattcgat tcctgtttgt aattgtcctt ttaacagtga tcgcgtattt
      601 cgtctcgctc aggcgcaatc acgaatgaat aacggtttgg ttgatgcgag tgattttgat
      661 gacgagcgta atggctggcc tgttgaacaa gtctggaaag aaatgcataa acttttgcca
      721 ttctcaccgg attcagtcgt cactcatggt gatttctcac ttgataacct tatttttgac
      781 gaggggaaat taataggttg tattgatgtt ggacgagtcg gaatcgcaga ccgataccag
      841 gatcttgcca tcctatggaa ctgcctcggt gagttttctc cttcattaca gaaacggctt
      901 tttcaaaaat atggtattga taatcctgat atgaataaat tgcagtttca tttgatgctc
      961 gatgagtttt tctaagaatt aattcatgag cggatacata tttgaatgta tttagaaaaa
     1021 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctaaat tgtaagcgtt
     1081 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     1141 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt
     1201 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga
     1261 aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg
     1321 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct
     1381 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc
     1441 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt
     1501 aatgcgccgc tacagggcgc gtcccattcg ccaatccgga tatagttcct cctttcagca
     1561 aaaaacccct caagacccgt ttagaggccc caaggggtta tgctagttat tgctcagcgg
     1621 tggcagcagc caactcagct tcctttcggg ctttgtttag cagcctaggt taattaagcc
     1681 tcgagagcag cagaagtaga gctgtccatg tgctggcgtt cgaatttagc agcagcggtt
     1741 tctttactac cgcgtggcac cagagcgagc tctgcggccg caagcttgtc gacggacgtc
     1801 gggcgcgcca aggcctgtac agaattcgga tccttaatgg tgatggtggt gatggggtcc
     1861 ctgaaagagg acttcaagag ccgcatttaa ccgacgacgt aggtcgtcca attcctggtc
     1921 aatcaccgca tcatcttgac tggcattatt agctggtact gaggaatcaa caggagccgc
     1981 ttcgagttgg ggcgtagcgg cagaggttcc cggtaatgct ccaccggcca tggaagcttt
     2041 cagggcggcc aactcgtctt ctaccccact gctggcttcc aactgggcaa actggttctc
     2101 gatgccaaag ccggctaact cccctgccgc ttgggaagtg gcttccatat ccagtacctt
     2161 gttctccatc cgttcaaaag cactagtagc actgctagta cctaagcccc cgagggtttg
     2221 ctgcagttca gcattagcct tggccgcttt ggccctggcc tgcaacatat ttttcttggt
     2281 tttagcttcg gaaatttttg cttctagagc cgcgaggttg cgacggagat tctccgacat
     2341 ggtcctctgt tgggctagct gggtttggta cgccgccgcc gtatctgtca gacttttttt
     2401 gcgggccagg gcttccctag ccaaattttc ttccccattg gtgagggcta attttgcccg
     2461 gtcttcccat ttcttggctt cctgggtgtc ttggttaaga cgttgctctg ttcgtttttc
     2521 ttcggcgatc gtgcgggcaa cggcctggcg gagttgcacc aggtcttcct gcatatcgat
     2581 cacggcctgt tccagaactt tttctggatc catatgtata tctccttctt aaagttaaac
     2641 aaaattattt ctagagggga attgttatcc gctcacaatt cccctatagt gagtcgtatt
     2701 aatttcgcgg gatcgagatc gatctcgatc ctctacgccg gacgcatcgt ggccggcatc
     2761 accggcgcca caggtgcggt tgctggcgcc tatatcgccg acatcaccga tggggaagat
     2821 cgggctcgcc acttcgggct catgagcgct tgtttcggcg tgggtatggt ggcaggcccc
     2881 gtggccgggg gactgttggg cgccatctcc ttgcatgcac cattccttgc ggcggcggtg
     2941 ctcaacggcc tcaacctact actgggctgc ttcctaatgc aggagtcgca taagggagag
     3001 cgtcgagatc ccggacacca tcgaatggcg caaaaccttt cgcggtatgg catgatagcg
     3061 cccggaagag agtcaattca gggtggtgaa tgtgaaacca gtaacgttat acgatgtcgc
     3121 agagtatgcc ggtgtctctt atcagaccgt ttcccgcgtg gtaaaccagg ccagccacgt
     3181 ttctgcgaaa acgcgggaaa aagtggaagc ggcgatggcg gagctgaatt acattcccaa
     3241 ccgcgtggca caacaactgg cgggcaaaca gtcgttgctg attggcgtta ccacctccag
     3301 tctggccctg cacgcgccgt cgcaaattgt cgcggcgatt aaatctcgcg ccgatcaact
     3361 gggtgccagc gtggtggtgt cgatggtaga acgaagcggc gtcgaagcct gtaaagcggc
     3421 ggtgcacaat cttctcgcgc aacgcgtcag tgggctgatc attaactatc cgctggatga
     3481 ccaggatgcc attgctgtgg aagctgcctg cactaatgtt ccggcgttat ttcttgatgt
     3541 ctctgaccag acacccatca acagtattat tttctcccat gaagacggta cgcgactggg
     3601 cgtggagcat ctggtcgcat tgggtcacca gcaaatcgcg ctgttagcgg gcccattaag
     3661 ttctgtctcg gcgcgtctgc gtctggctgg ctggcataaa tatctcactc gcaatcaaat
     3721 tcagccgata gcggaacggg aaggcgactg gagtgccatg tccggttttc aacaaaccat
     3781 gcaaatgctg aatgagggca tcgttcccac tgcgatgctg gttgccaacg atcagatggc
     3841 gctgggcgca atgcgcgcca ttaccgagtc cgggctgcgc gttggtgcgg acatctcggt
     3901 agtgggatac gacgataccg aagacagctc atgttatatc ccgccgttaa ccaccatcaa
     3961 acaggatttt cgcctgctgg ggcaaaccag cgtggaccgc ttgctgcaac tctctcaggg
     4021 ccaggcggtg aagggcaatc agctgttgcc cgtctcactg gtgaaaagaa aaaccaccct
     4081 ggcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
     4141 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtaagttagc
     4201 tcactcatta ggcaccggga tctcgaccga tgcccttgag agccttcaac ccagtcagct
     4261 ccttccggtg ggcgcggggc atgactagca tgatcgtgct cctgtcgttg aggacccggc
     4321 taggctggcg gggttgcctt actggttagc agaatgaatc accgatacgc gagcgaacgt
     4381 gaagcgactg ctgctgcaaa acgtctgcga cctgagcaac aacatgaatg gtcttcggtt
     4441 tccgtgtttc gtaaagtctg gaaacgcgga agtcagcgcc ctgcaccatt atgttccgga
     4501 tctgcatcgc aggatgctgc tggctaccct gtggaacacc tacatctgta ttaacgaagc
     4561 gctggcattg accctgagtg atttttctct ggtcccgccg catccatacc gccagttgtt
     4621 taccctcaca acgttccagt aaccgggcat gttcatcatc agtaacccgt atcgtgagca
     4681 tcctctctcg tttcatcggt atcattaccc ccatgaacag aaatccccct tacacggagg
     4741 catcagtgac caaacaggaa aaaaccgccc ttaacatggc ccgctttatc agaagccaga
     4801 cattaacgct tctggagaaa ctcaacgagc tggacgcgga tgaacaggca gacatctgtg
     4861 aatcgcttca cgaccacgct gatgagcttt accgcagctg cctcgcgcgt ttcggtgatg
     4921 acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg
     4981 atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg
     5041 cagccatgac ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc
     5101 agagcagatt gtactgagag tgcaccattg cggtgtgaaa taccgcacag atgcgtaagg
     5161 agaaaatacc gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     5221 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     5281 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     5341 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     5401 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     5461 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     5521 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     5581 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     5641 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     5701 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     5761 acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc
     5821 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     5881 caaaccaccg ctggtagcgg tggttttttt gttt
//
