LOCUS       sequence-274485-        5757 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  T7-tagged streptavidin with native C-terminus controlled by PT7 
            promoter; KanR, pBR322 ori.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5757)
  AUTHORS   Jeschek M, Bahls MO, Schneider V, Marliere P, Ward TR, Panke S
  TITLE     Biotin-independent strains of Escherichia coli for enhanced 
            streptavidin production.
  JOURNAL   Metab Eng. 2017 Mar;40:33-40. doi: 10.1016/j.ymben.2016.12.013. Epub
            2017 Jan 3.
  PUBMED    28062280
REFERENCE   2  (bases 1 to 5757)
  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: "Metab 
            Eng."; date: "2017-03"; volume: "40:33-40. doi"; pages: " 
            10.1016/j.ymben.2016.12.013. Epub 2017 Jan 3"
COMMENT     Sequence Label: pET30b_T7Sav
FEATURES             Location/Qualifiers
     source          1..5757
                     /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             89..121
                     /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             618..635
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(688..706)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      702..749
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      786..1241
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(873..892)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1083..1104
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1333..2148)
                     /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(1333..2148)
                     /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     2056..2075
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2140..2159
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2270..2858
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2759..2778
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3012..3029
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3044..3183
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3109..3128)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3269..3291
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3285..3476)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4285..5367)
                     /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     5329..5348
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5368..5445)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5651..5670
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat ggctagcatg actggtggac agcaaatggg
      121 tcgggatcag gccggcatca ccggcacctg gtacaaccag ctcggctcga ccttcatcgt
      181 gaccgcgggc gccgacggcg ccctgaccgg aacctacgag tcggccgtcg gcaacgccga
      241 gagccgctac gtcctgaccg gtcgttacga cagcgccccg gccaccgacg gcagcggcac
      301 cgccctcggt tggacggtgg cctggaagaa taactaccgc aacgcccact ccgcgaccac
      361 gtggagcggc cagtacgtcg gcggcgccga ggcgaggatc aacacccagt ggctgctgac
      421 ctccggcacc accgaggcca acgcctggaa gtccacgctg gtcggccacg acaccttcac
      481 caaggtgaag ccgtccgccg cctccatcga cgcggcgaag aaggccggcg tcaacaacgg
      541 caacccgctc gacgccgttc agcagtaata aggatccgaa ttcgagctcc gtcgacaagc
      601 ttgcggccgc actcgagcac caccaccacc accactgaga tccggctgct aacaaagccc
      661 gaaaggaagc tgagttggct gctgccaccg ctgagcaata actagcataa ccccttgggg
      721 cctctaaacg ggtcttgagg ggttttttgc tgaaaggagg aactatatcc ggattggcga
      781 atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
      841 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
      901 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
      961 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag
     1021 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     1081 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga
     1141 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
     1201 atttaacgcg aattttaaca aaatattaac gcttacaatt taggtggcac ttttcgggga
     1261 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     1321 atgaattaat tcttagaaaa actcatcgag catcaaatga aactgcaatt tattcatatc
     1381 aggattatca ataccatatt tttgaaaaag ccgtttctgt aatgaaggag aaaactcacc
     1441 gaggcagttc cataggatgg caagatcctg gtatcggtct gcgattccga ctcgtccaac
     1501 atcaatacaa cctattaatt tcccctcgtc aaaaataagg ttatcaagtg agaaatcacc
     1561 atgagtgacg actgaatccg gtgagaatgg caaaagttta tgcatttctt tccagacttg
     1621 ttcaacaggc cagccattac gctcgtcatc aaaatcactc gcatcaacca aaccgttatt
     1681 cattcgtgat tgcgcctgag cgagacgaaa tacgcgatcg ctgttaaaag gacaattaca
     1741 aacaggaatc gaatgcaacc ggcgcaggaa cactgccagc gcatcaacaa tattttcacc
     1801 tgaatcagga tattcttcta atacctggaa tgctgttttc ccggggatcg cagtggtgag
     1861 taaccatgca tcatcaggag tacggataaa atgcttgatg gtcggaagag gcataaattc
     1921 cgtcagccag tttagtctga ccatctcatc tgtaacatca ttggcaacgc tacctttgcc
     1981 atgtttcaga aacaactctg gcgcatcggg cttcccatac aatcgataga ttgtcgcacc
     2041 tgattgcccg acattatcgc gagcccattt atacccatat aaatcagcat ccatgttgga
     2101 atttaatcgc ggcctagagc aagacgtttc ccgttgaata tggctcataa caccccttgt
     2161 attactgttt atgtaagcag acagttttat tgttcatgac caaaatccct taacgtgagt
     2221 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     2281 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     2341 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     2401 agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     2461 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     2521 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     2581 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     2641 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     2701 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     2761 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     2821 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     2881 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     2941 attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa
     3001 cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc
     3061 tccttacgca tctgtgcggt atttcacacc gcaatggtgc actctcagta caatctgctc
     3121 tgatgccgca tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct
     3181 gcgccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca
     3241 tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg
     3301 tcatcaccga aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat
     3361 tcacagatgt ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat
     3421 gtctggcttc tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat
     3481 gcctccgtgt aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg
     3541 atgctcacga tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt
     3601 aaacaactgg cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag
     3661 cgcttcgtta atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag
     3721 atccggaaca taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgg
     3781 aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt
     3841 cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct
     3901 agccgggtcc tcaacgacag gagcacgatc atgcgcaccc gtggggccgc catgccggcg
     3961 ataatggcct gcttctcgcc gaaacgtttg gtggcgggac cagtgacgaa ggcttgagcg
     4021 agggcgtgca agattccgaa taccgcaagc gacaggccga tcatcgtcgc gctccagcga
     4081 aagcggtcct cgccgaaaat gacccagagc gctgccggca cctgtcctac gagttgcatg
     4141 ataaagaaga cagtcataag tgcggcgacg atagtcatgc cccgcgccca ccggaaggag
     4201 ctgactgggt tgaaggctct caagggcatc ggtcgagatc ccggtgccta atgagtgagc
     4261 taacttacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     4321 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag
     4381 ggtggttttt cttttcacca gtgagacggg caacagctga ttgcccttca ccgcctggcc
     4441 ctgagagagt tgcagcaagc ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt
     4501 gatggtggtt aacggcggga tataacatga gctgtcttcg gtatcgtcgt atcccactac
     4561 cgagatgtcc gcaccaacgc gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc
     4621 catctgatcg ttggcaacca gcatcgcagt gggaacgatg ccctcattca gcatttgcat
     4681 ggtttgttga aaaccggaca tggcactcca gtcgccttcc cgttccgcta tcggctgaat
     4741 ttgattgcga gtgagatatt tatgccagcc agccagacgc agacgcgccg agacagaact
     4801 taatgggccc gctaacagcg cgatttgctg gtgacccaat gcgaccagat gctccacgcc
     4861 cagtcgcgta ccgtcttcat gggagaaaat aatactgttg atgggtgtct ggtcagagac
     4921 atcaagaaat aacgccggaa cattagtgca ggcagcttcc acagcaatgg catcctggtc
     4981 atccagcgga tagttaatga tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc
     5041 cgctttacag gcttcgacgc cgcttcgttc taccatcgac accaccacgc tggcacccag
     5101 ttgatcggcg cgagatttaa tcgccgcgac aatttgcgac ggcgcgtgca gggccagact
     5161 ggaggtggca acgccaatca gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt
     5221 gggaatgtaa ttcagctccg ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac
     5281 gtggctggcc tggttcacca cgcgggaaac ggtctgataa gagacaccgg catactctgc
     5341 gacatcgtat aacgttactg gtttcacatt caccaccctg aattgactct cttccgggcg
     5401 ctatcatgcc ataccgcgaa aggttttgcg ccattcgatg gtgtccggga tctcgacgct
     5461 ctcccttatg cgactcctgc attaggaagc agcccagtag taggttgagg ccgttgagca
     5521 ccgccgccgc aaggaatggt gcatgcaagg agatggcgcc caacagtccc ccggccacgg
     5581 ggcctgccac catacccacg ccgaaacaag cgctcatgag cccgaagtgg cgagcccgat
     5641 cttccccatc ggtgatgtcg gcgatatagg cgccagcaac cgcacctgtg gcgccggtga
     5701 tgccggccac gatgcgtccg gcgtagagga tcgagatcga tctcgatccc gcgaaat
//
