LOCUS       sequence-322594-        6142 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses N-terminal (His)6-TEV cleavage site fusion of human gene 
            or portion of gene in bacterial strains. pET based vector..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6142)
  TITLE     Davis Lab plasmids
REFERENCE   2  (bases 1 to 6142)
  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: NKAP.001.140
FEATURES             Location/Qualifiers
     source          1..6142
                     /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             91..108
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             121..141
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     primer_bind     complement(643..661)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      657..704
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(879..884,885..901,902..907))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(934..952)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1020..1124
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1125..1193,1194..1985)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1343..1362)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2156..2744
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2645..2664
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2898..2915
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2930..3069
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2995..3014)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3155..3177
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3171..3362)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4674..5756)
                     /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     5718..5737
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5757..5834)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6040..6059
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atataccatg catcatcatc atcatcacag cagcggcaga
      121 gaaaacttgt atttccaggg catggctccg gtgtccggct cacgcagccc ggatagggag
      181 gcctcgggct cggggggaag acgtcgcagt tcgtcgaaga gtccgaagcc cagcaaatct
      241 gcccgctccc cgcggggccg ccgctctcgc tcgcactctt gctctcggtc cggggaccgg
      301 aatggactca cccatcagct gggtggcctc agccaaggct cccgaaacca gtcctaccgc
      361 tcacgctcgc ggtcgcgttc tagagagcgg ccctctgcgc cccggggcat ccccttcgct
      421 tctgcctcct cgtcagtcta ttacggcagc tactcgcgcc cctacgggag cgacaagcct
      481 tggcctagcc tcctcgacaa ggagagggag gagagcctgc ggcagaagag attaagtgag
      541 agagagagaa ttggagaatt gtagtgatga cgaagcttgc ggccgcactc gaggatccgg
      601 ctgctaacaa agcccgaaag gaagctgagt tggctgctgc caccgctgag caataactag
      661 cataacccct tggggcctct aaacgggtct tgaggggttt tttgctgaaa ggaggaacta
      721 tatccggata tcccgcaaga ggcccggcag taccggcata accaagccta tgcctacagc
      781 atccagggtg acggtgccga ggatgacgat gagcgcattg ttagatttca tacacggtgc
      841 ctgactgcgt tagcaattta actgtgataa actaccgcat taaagcttat cgatgataag
      901 ctgtcaaaca tgagaattct tgaagacgaa agggcctcgt gatacgccta tttttatagg
      961 ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc
     1021 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     1081 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     1141 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     1201 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     1261 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     1321 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtgtt gacgccgggc
     1381 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     1441 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     1501 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     1561 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     1621 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgca gcaatggcaa
     1681 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     1741 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     1801 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     1861 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     1921 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     1981 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     2041 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     2101 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     2161 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     2221 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     2281 gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga
     2341 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     2401 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     2461 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     2521 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     2581 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     2641 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     2701 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     2761 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
     2821 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca
     2881 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt
     2941 attttctcct tacgcatctg tgcggtattt cacaccgcaa tggtgcactc tcagtacaat
     3001 ctgctctgat gccgcatagt taagccagta tacactccgc tatcgctacg tgactgggtc
     3061 atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     3121 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     3181 tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtga
     3241 agcgattcac agatgtctgc ctgttcatcc gcgtccagct cgttgagttt ctccagaagc
     3301 gttaatgtct ggcttctgat aaagcgggcc atgttaaggg cggttttttc ctgtttggtc
     3361 actgatgcct ccgtgtaagg gggatttctg ttcatggggg taatgatacc gatgaaacga
     3421 gagaggatgc tcacgatacg ggttactgat gatgaacatg cccggttact ggaacgttgt
     3481 gagggtaaac aactggcggt atggatgcgg cgggaccaga gaaaaatcac tcagggtcaa
     3541 tgccagcgct tcgttaatac agatgtaggt gttccacagg gtagccagca gcatcctgcg
     3601 atgcagatcc ggaacataat ggtgcagggc gctgacttcc gcgtttccag actttacgaa
     3661 acacggaaac cgaagaccat tcatgttgtt gctcaggtcg cagacgtttt gcagcagcag
     3721 tcgcttcacg ttcgctcgcg tatcggtgat tcattctgct aaccagtaag gcaaccccgc
     3781 cagcctagcc gggtcctcaa cgacaggagc acgatcatgc gcacccgtgg ccaggaccca
     3841 acgctgcccg agatgcgccg cgtgcggctg ctggagatgg cggacgcgat ggatatgttc
     3901 tgccaagggt tggtttgcgc attcacagtt ctccgcaaga attgattggc tccaattctt
     3961 ggagtggtga atccgttagc gaggtgccgc cggcttccat tcaggtcgag gtggcccggc
     4021 tccatgcacc gcgacgcaac gcggggaggc agacaaggta tagggcggcg cctacaatcc
     4081 atgccaaccc gttccatgtg ctcgccgagg cggcataaat cgccgtgacg atcagcggtc
     4141 caatgatcga agttaggctg gtaagagccg cgagcgatcc ttgaagctgt ccctgatggt
     4201 cgtcatctac ctgcctggac agcatggcct gcaacgcggg catcccgatg ccgccggaag
     4261 cgagaagaat cataatgggg aaggccatcc agcctcgcgt cgcgaacgcc agcaagacgt
     4321 agcccagcgc gtcggccgcc atgccggcga taatggcctg cttctcgccg aaacgtttgg
     4381 tggcgggacc agtgacgaag gcttgagcga gggcgtgcaa gattccgaat accgcaagcg
     4441 acaggccgat catcgtcgcg ctccagcgaa agcggtcctc gccgaaaatg acccagagcg
     4501 ctgccggcac ctgtcctacg agttgcatga taaagaagac agtcataagt gcggcgacga
     4561 tagtcatgcc ccgcgcccac cggaaggagc tgactgggtt gaaggctctc aagggcatcg
     4621 gtcgagatcc cggtgcctaa tgagtgagct aacttacatt aattgcgttg cgctcactgc
     4681 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     4741 ggagaggcgg tttgcgtatt gggcgccagg gtggtttttc ttttcaccag tgagacgggc
     4801 aacagctgat tgcccttcac cgcctggccc tgagagagtt gcagcaagcg gtccacgctg
     4861 gtttgcccca gcaggcgaaa atcctgtttg atggtggtta acggcgggat ataacatgag
     4921 ctgtcttcgg tatcgtcgta tcccactacc gagatatccg caccaacgcg cagcccggac
     4981 tcggtaatgg cgcgcattgc gcccagcgcc atctgatcgt tggcaaccag catcgcagtg
     5041 ggaacgatgc cctcattcag catttgcatg gtttgttgaa aaccggacat ggcactccag
     5101 tcgccttccc gttccgctat cggctgaatt tgattgcgag tgagatattt atgccagcca
     5161 gccagacgca gacgcgccga gacagaactt aatgggcccg ctaacagcgc gatttgctgg
     5221 tgacccaatg cgaccagatg ctccacgccc agtcgcgtac cgtcttcatg ggagaaaata
     5281 atactgttga tgggtgtctg gtcagagaca tcaagaaata acgccggaac attagtgcag
     5341 gcagcttcca cagcaatggc atcctggtca tccagcggat agttaatgat cagcccactg
     5401 acgcgttgcg cgagaagatt gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct
     5461 accatcgaca ccaccacgct ggcacccagt tgatcggcgc gagatttaat cgccgcgaca
     5521 atttgcgacg gcgcgtgcag ggccagactg gaggtggcaa cgccaatcag caacgactgt
     5581 ttgcccgcca gttgttgtgc cacgcggttg ggaatgtaat tcagctccgc catcgccgct
     5641 tccacttttt cccgcgtttt cgcagaaacg tggctggcct ggttcaccac gcgggaaacg
     5701 gtctgataag agacaccggc atactctgcg acatcgtata acgttactgg tttcacattc
     5761 accaccctga attgactctc ttccgggcgc tatcatgcca taccgcgaaa ggttttgcgc
     5821 cattcgatgg tgtccgggat ctcgacgctc tcccttatgc gactcctgca ttaggaagca
     5881 gcccagtagt aggttgaggc cgttgagcac cgccgccgca aggaatggtg catgcaagga
     5941 gatggcgccc aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc
     6001 gctcatgagc ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc
     6061 gccagcaacc gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggat
     6121 cgagatctcg atcccgcgaa at
//
