LOCUS       sequence-181702-        7984 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pET LIC cloning vector for Biotin-His6-MBP-N10-tev-yORF-tdTomato.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7984)
  TITLE     C-terminal fluorescent fusion vectors for E. coli expression
REFERENCE   2  (bases 1 to 7984)
  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"
COMMENT     Sequence Label: pET MBP tdTomato LIC cloning vector (MBP-tdTom...
FEATURES             Location/Qualifiers
     source          1..7984
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     41..60
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     144..163
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        144..162
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    163..187
                     /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             202..224
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             246..290
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             306..323
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             333..1433
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     1407..1430
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1488..1508
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2991..3008
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3061..3079)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3075..3122
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3159..3614
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3246..3265)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3456..3477
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3707..4522)
                     /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(3707..4522)
                     /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     4430..4449
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4514..4533
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4644..5232
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5133..5152
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5386..5403
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5418..5557
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5483..5502)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5643..5665
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5659..5850)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(6659..7741)
                     /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     7703..7722
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7742..7819)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 cgctcatgag cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg gcgatatagg
       61 cgccagcaac cgcacctgtg gcgccggtga tgccggccac gatgcgtccg gcgtagagga
      121 tcgagatctc gatcccgcga aattaatacg actcactata ggggaattgt gagcggataa
      181 caattcccct ctagaaataa ttttgtttaa ctttaagaag gagatatagc atgggttctt
      241 ctggaggcct gaacgatatt tttgaagcgc agaaaattga atggcatgaa tccatgggtt
      301 cttctcacca tcaccatcac cacggttctt ctatgaaaat cgaagaaggt aaactggtaa
      361 tctggattaa cggcgataaa ggctataacg gtctcgctga agtcggtaag aaattcgaga
      421 aagataccgg aattaaagtc accgttgagc atccggataa actggaagag aaattcccac
      481 aggttgcggc aactggcgat ggccctgaca ttatcttctg ggcacacgac cgctttggtg
      541 gctacgctca atctggcctg ttggctgaaa tcaccccgga caaagcgttc caggacaagc
      601 tgtatccgtt tacctgggat gccgtacgtt acaacggcaa gctgattgct tacccgatcg
      661 ctgttgaagc gttatcgctg atttataaca aagatctgct gccgaacccg ccaaaaacct
      721 gggaagagat cccggcgctg gataaagaac tgaaagcgaa aggtaagagc gcgctgatgt
      781 tcaacctgca agaaccgtac ttcacctggc cgctgattgc tgctgacggg ggttatgcgt
      841 tcaagtatga aaacggcaag tacgacatta aagacgtggg cgtggataac gctggcgcga
      901 aagcgggtct gaccttcctg gttgacctga ttaaaaacaa acacatgaat gcagacaccg
      961 attactccat cgcagaagct gcctttaata aaggcgaaac agcgatgacc atcaacggcc
     1021 cgtgggcatg gtccaacatc gacaccagca aagtgaatta tggtgtaacg gtactgccga
     1081 ccttcaaggg tcaaccatcc aaaccgttcg ttggcgtgct gagcgcaggt attaacgccg
     1141 ccagtccgaa caaagagctg gcaaaagagt tcctcgaaaa ctatctgctg actgatgaag
     1201 gtctggaagc ggttaataaa gacaaaccgc tgggtgccgt agcgctgaag tcttacgagg
     1261 aagagttggc gaaagatcca cgtattgccg ccactatgga aaacgcccag aaaggtgaaa
     1321 tcatgccgaa catcccgcag atgtccgctt tctggtatgc cgtgcgtact gcggtgatca
     1381 acgccgccag cggtcgtcag actgtcgatg aagccctgaa agacgcgcag actaattcga
     1441 gctcgaacaa caacaacaat aacaataaca acaacctcgg gatcgaggaa aacctgtact
     1501 tccaatccaa tattggtagt gggagcaacg gcagcagcgg atccgtgagt aagggcgagg
     1561 aagtgatcaa agagttcatg cggtttaagg tgagaatgga aggaagcatg aacggccacg
     1621 agttcgaaat tgagggagaa ggagagggac ggccctacga gggcacccag acagccaagc
     1681 tgaaagtgac aaagggcggg cctctgccat tcgcttggga catcctgagc ccacagttta
     1741 tgtacggctc caaggcctat gtgaaacatc cagctgacat tcccgattat aagaaactga
     1801 gcttccccga ggggtttaag tgggaaagag tgatgaactt cgaggacgga ggcctggtga
     1861 ctgtgaccca ggacagctcc ctgcaggatg ggaccctgat ctacaaggtg aaaatgagag
     1921 ggacaaattt tccccctgat ggacctgtga tgcagaagaa aactatggga tgggaggcct
     1981 ccaccgaaag gctgtatcca cgcgacgggg tgctgaaagg agaaatccac caggctctga
     2041 agctgaaaga tgggggacat tacctggtgg agttcaagac aatctacatg gccaagaaac
     2101 ctgtgcagct gccaggctac tattacgtgg acacaaaact ggatatcact tcacacaacg
     2161 aggactacac tattgtggag cagtatgaac ggagcgaggg gagacaccat ctgttcctgg
     2221 gccatgggac tggaagtacc ggctcagggt ctagtggaac cgcctcaagc gaggataaca
     2281 atatggctgt gatcaaagag ttcatgaggt ttaaggtgcg catggagggc agcatgaatg
     2341 ggcacgaatt tgagattgaa ggagagggcg aagggaggcc ttacgagggc acacagactg
     2401 ccaagctgaa agtgaccaag ggaggaccac tgcctttcgc ttgggatatc ctgtctcctc
     2461 agtttatgta cggaagtaag gcctatgtca agcatcccgc tgacattcct gattacaaga
     2521 aactgtcttt cccagagggc tttaagtggg agagagtgat gaattttgaa gatggaggcc
     2581 tggtgaccgt gacacaggac tcctctctgc aggatggcac tctgatctac aaagtcaaaa
     2641 tgcgcggcac caattttcca cccgatgggc ccgtgatgca gaagaaaaca atggggtggg
     2701 aggccagcac tgaacggctg tatcctagag acggagtgct gaagggcgaa atccaccagg
     2761 ccctgaagct gaaagacggc ggccactacc tggtggagtt caaaaccatc tacatggcca
     2821 agaaaccagt gcagctgccc ggctattact atgtggacac caagctggat atcacatccc
     2881 acaatgaaga ctacaccatt gtggaacagt atgagaggtc tgaaggacgc caccatctgt
     2941 ttctgtacgg catggatgag ctgtataagt aataagcggc cgcactcgag caccaccacc
     3001 accaccactg agatccggct gctaacaaag cccgaaagga agctgagttg gctgctgcca
     3061 ccgctgagca ataactagca taaccccttg gggcctctaa acgggtcttg aggggttttt
     3121 tgctgaaagg aggaactata tccggattgg cgaatgggac gcgccctgta gcggcgcatt
     3181 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     3241 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     3301 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     3361 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     3421 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     3481 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     3541 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaactagt
     3601 aacgtttaca atttcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     3661 atttttctaa atacattcaa atatgtatcc gctcatgaat taattcttag aaaaactcat
     3721 cgagcatcaa atgaaactgc aatttattca tatcaggatt atcaatacca tatttttgaa
     3781 aaagccgttt ctgtaatgaa ggagaaaact caccgaggca gttccatagg atggcaagat
     3841 cctggtatcg gtctgcgatt ccgactcgtc caacatcaat acaacctatt aatttcccct
     3901 cgtcaaaaat aaggttatca agtgagaaat caccatgagt gacgactgaa tccggtgaga
     3961 atggcaaaag tttatgcatt tctttccaga cttgttcaac aggccagcca ttacgctcgt
     4021 catcaaaatc actcgcatca accaaaccgt tattcattcg tgattgcgcc tgagcgagac
     4081 gaaatacgcg atcgctgtta aaaggacaat tacaaacagg aatcgaatgc aaccggcgca
     4141 ggaacactgc cagcgcatca acaatgtttt cacctgaatc aggatattct tctaatacct
     4201 ggaatgctgt tttcccgggg atcgcagtgg tgagtaacca tgcatcatca ggagtacgga
     4261 taaaatgctt gatggtcgga agaggcataa attccgtcag ccagtttagt ctgaccatct
     4321 catctgtaac atcattggca acgctacctt tgccatgttt cagaaacaac tctggcgcat
     4381 cgggcttccc atacaatcga tagattgtcg cacctgattg cccgacatta tcgcgagccc
     4441 atttataccc atataaatca gcatccatgt tggaatttaa tcgcggccta gagcaagacg
     4501 tttcccgttg aatatggctc ataacacccc ttgtattact gtttatgtaa gcagacagtt
     4561 ttattgttca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     4621 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     4681 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     4741 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg
     4801 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     4861 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     4921 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     4981 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     5041 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     5101 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     5161 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     5221 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     5281 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     5341 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     5401 gaggaagcgg aagagcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     5461 caccgcaatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtata
     5521 cactccgcta tcgctacgtg actgggtcat ggctgcgccc cgacacccgc caacacccgc
     5581 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt
     5641 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgaggcagct
     5701 gcggtaaagc tcatcagcgt ggtcgtgaag cgattcacag atgtctgcct gttcatccgc
     5761 gtccagctcg ttgagtttct ccagaagcgt taatgtctgg cttctgataa agcgggccat
     5821 gttaagggcg gttttttcct gtttggtcac tgatgcctcc gtgtaagggg gatttctgtt
     5881 catgggggta atgataccga tgaaacgaga gaggatgctc acgatacggg ttactgatga
     5941 tgaacatgcc cggttactgg aacgttgtga gggtaaacaa ctggcggtat ggatgcggcg
     6001 ggaccagaga aaaatcactc agggtcaatg ccagcgcttc gttaatacag atgtaggtgt
     6061 tccacagggt agccagcagc atcctgcgat gcagatccgg aacataatgg tgcagggcgc
     6121 tgacttccgc gtttccagac tttacgaaac acggaaaccg aagaccattc atgttgttgc
     6181 tcaggtcgca gacgttttgc agaagcagtc gcttcacgtt cgctcgcgta tcggtgattc
     6241 attctgctaa ccagtaaggc aaccccgcca gcctagccgg gtcctcaacg acaggagcac
     6301 gatcatgcgc acccgtgggg ccgccatgcc ggcgataatg gcctgcttct cgccgaaacg
     6361 tttggtggcg ggaccagtga cgaaggcttg agcgagggcg tgcaagattc cgaataccgc
     6421 aagcgacagg ccgatcatcg tcgcgctcca gcgaaagcgg tcctcgccga aaatgaccca
     6481 gagcgctgcc ggcacctgtc ctacgagttg catgataaag aagacagtca taagtgcggc
     6541 gacgatagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg ctctcaaggg
     6601 catcggtcga gatcccggtg cctaatgagt gagctaactt acattaattg cgttgcgctc
     6661 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     6721 cgcggggaga ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga
     6781 cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca
     6841 cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac
     6901 atgagctgtc ttcggtatcg tcgtatccca ctaccgagat atccgcacca acgcgcagcc
     6961 cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg
     7021 cagtgggaac gataccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac
     7081 tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc
     7141 agccagccag acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt
     7201 gctggtgacc caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga
     7261 aaataatact gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag
     7321 tgcaggcagc ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc
     7381 cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc
     7441 gttctaccat cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg
     7501 cgacaatttg cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg
     7561 actgtttgcc cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg
     7621 ccgcttccac tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg
     7681 aaacggtctg ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca
     7741 cattcaccac cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt
     7801 tgcgccattc gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg
     7861 aagcagccca gtagtaggtt gaggccgttg agcaccgccg ccgcaaggaa tggtgcatgc
     7921 aaggagatgg cgcccaacag tcccccggcc acggggcctg ccaccatacc cacgccgaaa
     7981 caag
//
