LOCUS       sequence-461309-        5783 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression for structure determination; may not be full 
            ORF.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5783)
  TITLE     Structural Genomics Consortium Clones
REFERENCE   2  (bases 1 to 5783)
  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: UBC32
FEATURES             Location/Qualifiers
     source          1..5783
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(119..574)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(256..277)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     468..487
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(611..658)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     654..672
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(725..742)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1246..1263)
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             complement(1273..1290)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             1309..1331
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1346..1370
                     /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)."
     primer_bind     complement(1370..1389)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1371..1389)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1473..1492)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1698..1775
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1776..2858
                     /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     complement(1795..1814)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3667..3858
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3852..3874)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3960..4099
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4015..4034
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4114..4131)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4285..4873)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4365..4384)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4984..5003)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             join(4995..5783,1..27)
                     /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             join(4995..5783,1..27)
                     /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     complement(5068..5087)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 catttgatgc tcgatgagtt tttctaagaa ttaattcatg agcggataca tatttgaatg
       61 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctaa
      121 attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat cagctcattt
      181 tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata gaccgagata
      241 gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt ggactccaac
      301 gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc atcaccctaa
      361 tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa agggagcccc
      421 cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg gaagaaagcg
      481 aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt aaccaccaca
      541 cccgccgcgc ttaatgcgcc gctacagggc gcgtcccatt cgccaatccg gatatagttc
      601 ctcctttcag caaaaaaccc ctcaagaccc gtttagaggc cccaaggggt tatgctagtt
      661 attgctcagc ggtggcagca gccaactcag cttcctttcg ggctttgtta gcagccggat
      721 ctcagtggtg gtggtggtgg tgctcgagtg cggccgcaag cttcgtcatc atgagggcgc
      781 aaacttcttg gcttgtgctc ggaccctttt ctcgtactcc actctgtttt ggcagtaaat
      841 cgtgtaggcc tctgcttgag ctgggtcttg gatatttggt tcatttagaa gttcctgtat
      901 tcctaatagg atctgtttga ttgtgatggc tggcctccag tccttgtcct cctctaagat
      961 ggacaggcac actgtccccg aagggtacac attcgggtga aataatggtg gttcgaattt
     1021 acattttggt ggcgaagatg gataatcatc tttgaaaagc atccgtagtt taaacaagcc
     1081 tccttcccac ggagtccctt tctttcctgg aatggcgcac tcccagttca tgaggttcat
     1141 cgtgccatcg ggattttttg ttgggacagc cacgaaacca aatgggtggt ctttcctcca
     1201 tgctttcctc tcctgggcga gtctgctgag ggcgatcccc gacatactac cacgcggaac
     1261 caggccgctg ctgtgatgat gatgatgatg gctgctgccc atggtatatc tccttcttaa
     1321 agttaaacaa aattatttct agaggggaat tgttatccgc tcacaattcc cctatagtga
     1381 gtcgtattaa tttcgcggga tcgagatctc gatcctctac gccggacgca tcgtggccgg
     1441 catcaccggc gccacaggtg cggttgctgg cgcctatatc gccgacatca ccgatgggga
     1501 agatcgggct cgccacttcg ggctcatgag cgcttgtttc ggcgtgggta tggtggcagg
     1561 ccccgtggcc gggggactgt tgggcgccat ctccttgcat gcaccattcc ttgcggcggc
     1621 ggtgctcaac ggcctcaacc tactactggg ctgcttccta atgcaggagt cgcataaggg
     1681 agagcgtcga gatcccggac accatcgaat ggcgcaaaac ctttcgcggt atggcatgat
     1741 agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg
     1801 tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac caggccagcc
     1861 acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg aattacattc
     1921 ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt gctgattggc gttgccacct
     1981 ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc
     2041 aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag
     2101 cggcggtgca caatcttctc gcgcaacgcg tcagtgggct gatcattaac tatccgctgg
     2161 atgaccagga tgccattgct gtggaagctg cctgcactaa tgttccggcg ttatttcttg
     2221 atgtctctga ccagacaccc atcaacagta ttattttctc ccatgaagac ggtacgcgac
     2281 tgggcgtgga gcatctggtc gcattgggtc accagcaaat cgcgctgtta gcgggcccat
     2341 taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcgcaatc
     2401 aaattcagcc gatagcggaa cgggaaggcg actggagtgc catgtccggt tttcaacaaa
     2461 ccatgcaaat gctgaatgag ggcatcgttc ccactgcgat gctggttgcc aacgatcaga
     2521 tggcgctggg cgcaatgcgc gccattaccg agtccgggct gcgcgttggt gcggatatct
     2581 cggtagtggg atacgacgat accgaagaca gctcatgtta tatcccgccg ttaaccacca
     2641 tcaaacagga ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg caactctctc
     2701 agggccaggc ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa agaaaaacca
     2761 ccctggcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc
     2821 tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtaagt
     2881 tagctcactc attaggcacc gggatctcga ccgatgccct tgagagcctt caacccagtc
     2941 agctccttcc ggtgggcgcg gggcatgact atcgtcgccg cacttatgac tgtcttcttt
     3001 atcatgcaac tcgtaggaca ggtgccggca gcgctctggg tcattttcgg cgaggaccgc
     3061 tttcgctgga gcgcgacgat gatcggcctg tcgcttgcgg tattcggaat cttgcacgcc
     3121 ctcgctcaag ccttcgtcac tggtcccgcc accaaacgtt tcggcgagaa gcaggccatt
     3181 atcgccggca tggcggcccc acgggtgcgc atgatcgtgc tcctgtcgtt gaggacccgg
     3241 ctaggctggc ggggttgcct tactggttag cagaatgaat caccgatacg cgagcgaacg
     3301 tgaagcgact gctgctgcaa aacgtctgcg acctgagcaa caacatgaat ggtcttcggt
     3361 ttccgtgttt cgtaaagtct ggaaacgcgg aagtcagcgc cctgcaccat tatgttccgg
     3421 atctgcatcg caggatgctg ctggctaccc tgtggaacac ctacatctgt attaacgaag
     3481 cgctggcatt gaccctgagt gatttttctc tggtcccgcc gcatccatac cgccagttgt
     3541 ttaccctcac aacgttccag taaccgggca tgttcatcat cagtaacccg tatcgtgagc
     3601 atcctctctc gtttcatcgg tatcattacc cccatgaaca gaaatccccc ttacacggag
     3661 gcatcagtga ccaaacagga aaaaaccgcc cttaacatgg cccgctttat cagaagccag
     3721 acattaacgc ttctggagaa actcaacgag ctggacgcgg atgaacaggc agacatctgt
     3781 gaatcgcttc acgaccacgc tgatgagctt taccgcagct gcctcgcgcg tttcggtgat
     3841 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
     3901 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
     3961 gcagccatga cccagtcacg tagcgatagc ggagtgtata ctggcttaac tatgcggcat
     4021 cagagcagat tgtactgaga gtgcaccatt gcggtgtgaa ataccgcaca gatgcgtaag
     4081 gagaaaatac cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     4141 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     4201 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     4261 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     4321 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     4381 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     4441 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     4501 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     4561 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     4621 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     4681 tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat
     4741 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     4801 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     4861 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     4921 aaactcacgt taagggattt tggtcatgaa caataaaact gtctgcttac ataaacagta
     4981 atacaagggg tgttatgagc catattcaac gggaaacgtc ttgctctagg ccgcgattaa
     5041 attccaacat ggatgctgat ttatatgggt ataaatgggc tcgcgataat gtcgggcaat
     5101 caggtgcgac aatctatcga ttgtatggga agcccgatgc gccagagttg tttctgaaac
     5161 atggcaaagg tagcgttgcc aatgatgtta cagatgagat ggtcagacta aactggctga
     5221 cggaatttat gcctcttccg accatcaagc attttatccg tactcctgat gatgcatggt
     5281 tactcaccac tgcgatcccc gggaaaacag cattccaggt attagaagaa tatcctgatt
     5341 caggtgaaaa tattgttgat gcgctggcag tgttcctgcg ccggttgcat tcgattcctg
     5401 tttgtaattg tccttttaac agcgatcgcg tatttcgtct cgctcaggcg caatcacgaa
     5461 tgaataacgg tttggttgat gcgagtgatt ttgatgacga gcgtaatggc tggcctgttg
     5521 aacaagtctg gaaagaaatg cataaacttt tgccattctc accggattca gtcgtcactc
     5581 atggtgattt ctcacttgat aaccttattt ttgacgaggg gaaattaata ggttgtattg
     5641 atgttggacg agtcggaatc gcagaccgat accaggatct tgccatccta tggaactgcc
     5701 tcggtgagtt ttctccttca ttacagaaac ggctttttca aaaatatggt attgataatc
     5761 ctgatatgaa taaattgcag ttt
//
