LOCUS       sequence-296541-        6130 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression/crystallography (PDB:5QT0).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6130)
  TITLE     Opher Gileadi SGC expression vectors
REFERENCE   2  (bases 1 to 6130)
  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-c021
FEATURES             Location/Qualifiers
     source          1..6130
                     /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             101..118
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             950..994
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             995..1012
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1065..1083)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1079..1126
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1163..1618
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1250..1269)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1460..1481
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1710..2525)
                     /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(1710..2525)
                     /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     2433..2452
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2517..2536
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2647..3235
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3136..3155
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3389..3406
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3421..3560
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3486..3505)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3646..3668
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3662..3853)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4662..5744)
                     /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     5706..5725
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5745..5822)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6028..6047
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gtgcagcagc catcatcatc atcatcacgg
      121 cagcggcagc ggctctgacc aggaggcaaa accttcaact gaggacttgg gggataagaa
      181 ggaaggtgaa tatattaaac tcaaagtcat tggacaggat agcagtgaga ttcacttcaa
      241 agtgaaaatg acaacacatc tcaagaaact caaagaatca tactgtcaaa gacagggtgt
      301 tccaatgaat tcactcaggt ttctctttga gggtcagaga attgctgata atcatactcc
      361 aaaagaactg ggaatggagg aagaagatgt gattgaagtt taccaggagc aaacgggagg
      421 tggagaactg cgtgtgggtc tggaagaaag cgaactgtgg ctgcgcttta aagagctgac
      481 caacgaaatg atcgtgacca aaaacggccg ccgcatgttt cctgtgctga aagttaacgt
      541 gagcggcctg gacccgaacg ccatgtacag cttcctgctg gatttcgttg ccgcagataa
      601 ccaccgctgg aagtatgtga acggcgaatg ggtgccgggt ggcaaaccgg aaccgcaggc
      661 acctagctgc gtgtatatcc acccggatag ccctaacttc ggtgcccatt ggatgaaagc
      721 cccggtgagc ttcagcaagg tgaagctgac caacaaactg aacggcggcg gccaaatcat
      781 gctgaacagc ttacataaat atgaacctcg catccacatt gtgcgtgtgg gcgacccgca
      841 acgtatgatc accagccatt gcttcccgga gacacagttc attgccgtta ccgcctacca
      901 gaacgaggaa attaccgcac tgaaaattaa atacaactga agcggtggcg gtctgaacga
      961 catcttcgag gctcagaaaa tcgaatggca cgaacaccac caccaccacc actgagatcc
     1021 ggctgctaac aaagcccgaa aggaagctga gttggctgct gccaccgctg agcaataact
     1081 agcataaccc cttggggcct ctaaacgggt cttgaggggt tttttgctga aaggaggaac
     1141 tatatccgga ttggcgaatg ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     1201 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct
     1261 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     1321 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag
     1381 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     1441 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc
     1501 tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat
     1561 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttag
     1621 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     1681 caaatatgta tccgctcatg aattaattct tagaaaaact catcgagcat caaatgaaac
     1741 tgcaatttat tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat
     1801 gaaggagaaa actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg
     1861 attccgactc gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta
     1921 tcaagtgaga aatcaccatg agtgacgact gaatccggtg agaatggcaa aagtttatgc
     1981 atttctttcc agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca
     2041 tcaaccaaac cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg
     2101 ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca
     2161 tcaacaatat tttcacctga atcaggatat tcttctaata cctggaatgc tgttttcccg
     2221 gggatcgcag tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc
     2281 ggaagaggca taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg
     2341 gcaacgctac ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat
     2401 cgatagattg tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa
     2461 tcagcatcca tgttggaatt taatcgcggc ctagagcaag acgtttcccg ttgaatatgg
     2521 ctcataacac cccttgtatt actgtttatg taagcagaca gttttattgt tcatgaccaa
     2581 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     2641 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     2701 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     2761 tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca
     2821 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     2881 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     2941 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     3001 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     3061 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     3121 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     3181 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     3241 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     3301 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     3361 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
     3421 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca atggtgcact
     3481 ctcagtacaa tctgctctga tgccgcatag ttaagccagt atacactccg ctatcgctac
     3541 gtgactgggt catggctgcg ccccgacacc cgccaacacc cgctgacgcg ccctgacggg
     3601 cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg agctgcatgt
     3661 gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa agctcatcag
     3721 cgtggtcgtg aagcgattca cagatgtctg cctgttcatc cgcgtccagc tcgttgagtt
     3781 tctccagaag cgttaatgtc tggcttctga taaagcgggc catgttaagg gcggtttttt
     3841 cctgtttggt cactgatgcc tccgtgtaag ggggatttct gttcatgggg gtaatgatac
     3901 cgatgaaacg agagaggatg ctcacgatac gggttactga tgatgaacat gcccggttac
     3961 tggaacgttg tgagggtaaa caactggcgg tatggatgcg gcgggaccag agaaaaatca
     4021 ctcagggtca atgccagcgc ttcgttaata cagatgtagg tgttccacag ggtagccagc
     4081 agcatcctgc gatgcagatc cggaacataa tggtgcaggg cgctgacttc cgcgtttcca
     4141 gactttacga aacacggaaa ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt
     4201 tgcagcagca gtcgcttcac gttcgctcgc gtatcggtga ttcattctgc taaccagtaa
     4261 ggcaaccccg ccagcctagc cgggtcctca acgacaggag cacgatcatg cgcacccgtg
     4321 gggccgccat gccggcgata atggcctgct tctcgccgaa acgtttggtg gcgggaccag
     4381 tgacgaaggc ttgagcgagg gcgtgcaaga ttccgaatac cgcaagcgac aggccgatca
     4441 tcgtcgcgct ccagcgaaag cggtcctcgc cgaaaatgac ccagagcgct gccggcacct
     4501 gtcctacgag ttgcatgata aagaagacag tcataagtgc ggcgacgata gtcatgcccc
     4561 gcgcccaccg gaaggagctg actgggttga aggctctcaa gggcatcggt cgagatcccg
     4621 gtgcctaatg agtgagctaa cttacattaa ttgcgttgcg ctcactgccc gctttccagt
     4681 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     4741 tgcgtattgg gcgccagggt ggtttttctt ttcaccagtg agacgggcaa cagctgattg
     4801 cccttcaccg cctggccctg agagagttgc agcaagcggt ccacgctggt ttgccccagc
     4861 aggcgaaaat cctgtttgat ggtggttaac ggcgggatat aacatgagct gtcttcggta
     4921 tcgtcgtatc ccactaccga gatatccgca ccaacgcgca gcccggactc ggtaatggcg
     4981 cgcattgcgc ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc
     5041 tcattcagca tttgcatggt ttgttgaaaa ccggacatgg cactccagtc gccttcccgt
     5101 tccgctatcg gctgaatttg attgcgagtg agatatttat gccagccagc cagacgcaga
     5161 cgcgccgaga cagaacttaa tgggcccgct aacagcgcga tttgctggtg acccaatgcg
     5221 accagatgct ccacgcccag tcgcgtaccg tcttcatggg agaaaataat actgttgatg
     5281 ggtgtctggt cagagacatc aagaaataac gccggaacat tagtgcaggc agcttccaca
     5341 gcaatggcat cctggtcatc cagcggatag ttaatgatca gcccactgac gcgttgcgcg
     5401 agaagattgt gcaccgccgc tttacaggct tcgacgccgc ttcgttctac catcgacacc
     5461 accacgctgg cacccagttg atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc
     5521 gcgtgcaggg ccagactgga ggtggcaacg ccaatcagca acgactgttt gcccgccagt
     5581 tgttgtgcca cgcggttggg aatgtaattc agctccgcca tcgccgcttc cactttttcc
     5641 cgcgttttcg cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt ctgataagag
     5701 acaccggcat actctgcgac atcgtataac gttactggtt tcacattcac caccctgaat
     5761 tgactctctt ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg
     5821 tccgggatct cgacgctctc ccttatgcga ctcctgcatt aggaagcagc ccagtagtag
     5881 gttgaggccg ttgagcaccg ccgccgcaag gaatggtgca tgcaaggaga tggcgcccaa
     5941 cagtcccccg gccacggggc ctgccaccat acccacgccg aaacaagcgc tcatgagccc
     6001 gaagtggcga gcccgatctt ccccatcggt gatgtcggcg atataggcgc cagcaaccgc
     6061 acctgtggcg ccggtgatgc cggccacgat gcgtccggcg tagaggatcg agatctcgat
     6121 cccgcgaaat
//
