LOCUS       sequence-296505-        5851 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For protein expression/crystallography (PDB:6F58).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5851)
  TITLE     Opher Gileadi SGC expression vectors
REFERENCE   2  (bases 1 to 5851)
  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: TBXTA-c005
FEATURES             Location/Qualifiers
     source          1..5851
                     /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             646..663
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             716..733
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(786..804)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      800..847
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      884..1339
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(971..990)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1181..1202
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1431..2246)
                     /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(1431..2246)
                     /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     2154..2173
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2238..2257
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2368..2956
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2857..2876
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3110..3127
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3142..3281
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3207..3226)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3367..3389
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3383..3574)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4383..5465)
                     /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     5427..5446
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5466..5543)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5749..5768
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atataccatg ggcgaactgc gtgtgggtct ggaagaaagc
      121 gaactgtggc tgcgctttaa agagctgacc aacgaaatga tcgtgaccaa aaacggccgc
      181 cgcatgtttc ctgtgctgaa agttaacgtg agcggcctgg acccgaacgc catgtacagc
      241 ttcctgctgg atttcgttgc cgcagataac caccgctgga agtatgtgaa cggcgaatgg
      301 gtgccgggtg gcaaaccgga accgcaggca cctagctgcg tgtatatcca cccggatagc
      361 cctaacttcg gtgcccattg gatgaaagcc ccggtgagct tcagcaaggt gaagctgacc
      421 aacaaactga acggcggcgg ccaaatcatg ctgaacagct tacataaata tgaacctcgc
      481 atccacattg tgcgtgtggg cggcccgcaa cgtatgatca ccagccattg cttcccggag
      541 acacagttca ttgccgttac cgcctaccag aacgaggaaa ttaccgcact gaaaattaaa
      601 tacaaccctt ttgcaaaagc cttcttagat gccaaagagc gcagccacca ccaccatcat
      661 cactaatgag gatccgaatt cgagctccgt cgacaagctt gcggccgcac tcgagcacca
      721 ccaccaccac cactgagatc cggctgctaa caaagcccga aaggaagctg agttggctgc
      781 tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg tcttgagggg
      841 ttttttgctg aaaggaggaa ctatatccgg attggcgaat gggacgcgcc ctgtagcggc
      901 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
      961 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     1021 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     1081 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     1141 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     1201 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     1261 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     1321 atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     1381 gtttattttt ctaaatacat tcaaatatgt atccgctcat gaattaattc ttagaaaaac
     1441 tcatcgagca tcaaatgaaa ctgcaattta ttcatatcag gattatcaat accatatttt
     1501 tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga ggcagttcca taggatggca
     1561 agatcctggt atcggtctgc gattccgact cgtccaacat caatacaacc tattaatttc
     1621 ccctcgtcaa aaataaggtt atcaagtgag aaatcaccat gagtgacgac tgaatccggt
     1681 gagaatggca aaagtttatg catttctttc cagacttgtt caacaggcca gccattacgc
     1741 tcgtcatcaa aatcactcgc atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg
     1801 agacgaaata cgcgatcgct gttaaaagga caattacaaa caggaatcga atgcaaccgg
     1861 cgcaggaaca ctgccagcgc atcaacaata ttttcacctg aatcaggata ttcttctaat
     1921 acctggaatg ctgttttccc ggggatcgca gtggtgagta accatgcatc atcaggagta
     1981 cggataaaat gcttgatggt cggaagaggc ataaattccg tcagccagtt tagtctgacc
     2041 atctcatctg taacatcatt ggcaacgcta cctttgccat gtttcagaaa caactctggc
     2101 gcatcgggct tcccatacaa tcgatagatt gtcgcacctg attgcccgac attatcgcga
     2161 gcccatttat acccatataa atcagcatcc atgttggaat ttaatcgcgg cctagagcaa
     2221 gacgtttccc gttgaatatg gctcataaca ccccttgtat tactgtttat gtaagcagac
     2281 agttttattg ttcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     2341 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     2401 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     2461 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct
     2521 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     2581 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     2641 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     2701 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     2761 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     2821 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     2881 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     2941 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     3001 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     3061 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     3121 gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     3181 ttcacaccgc aatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     3241 tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac ccgccaacac
     3301 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     3361 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc
     3421 agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct gcctgttcat
     3481 ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg ataaagcggg
     3541 ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc
     3601 tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata cgggttactg
     3661 atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg gtatggatgc
     3721 ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat acagatgtag
     3781 gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata atggtgcagg
     3841 gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc attcatgttg
     3901 ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg
     3961 attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc aacgacagga
     4021 gcacgatcat gcgcacccgt ggggccgcca tgccggcgat aatggcctgc ttctcgccga
     4081 aacgtttggt ggcgggacca gtgacgaagg cttgagcgag ggcgtgcaag attccgaata
     4141 ccgcaagcga caggccgatc atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga
     4201 cccagagcgc tgccggcacc tgtcctacga gttgcatgat aaagaagaca gtcataagtg
     4261 cggcgacgat agtcatgccc cgcgcccacc ggaaggagct gactgggttg aaggctctca
     4321 agggcatcgg tcgagatccc ggtgcctaat gagtgagcta acttacatta attgcgttgc
     4381 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     4441 aacgcgcggg gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt
     4501 gagacgggca acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg
     4561 tccacgctgg tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata
     4621 taacatgagc tgtcttcggt atcgtcgtat cccactaccg agatatccgc accaacgcgc
     4681 agcccggact cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc
     4741 atcgcagtgg gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg
     4801 gcactccagt cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta
     4861 tgccagccag ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg
     4921 atttgctggt gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg
     4981 gagaaaataa tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca
     5041 ttagtgcagg cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc
     5101 agcccactga cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg
     5161 cttcgttcta ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc
     5221 gccgcgacaa tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc
     5281 aacgactgtt tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc
     5341 atcgccgctt ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg
     5401 cgggaaacgg tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt
     5461 ttcacattca ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag
     5521 gttttgcgcc attcgatggt gtccgggatc tcgacgctct cccttatgcg actcctgcat
     5581 taggaagcag cccagtagta ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc
     5641 atgcaaggag atggcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc
     5701 gaaacaagcg ctcatgagcc cgaagtggcg agcccgatct tccccatcgg tgatgtcggc
     5761 gatataggcg ccagcaaccg cacctgtggc gccggtgatg ccggccacga tgcgtccggc
     5821 gtagaggatc gagatctcga tcccgcgaaa t
//
