LOCUS       sequence-335161-        6219 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human TFIIB in bacteria. The resulting plasmid was used to
            generate a single expression construct encoding a His-tagged TFIIB.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6219)
  AUTHORS   He Y, Sato S, Tomomori-Sato C, Chen S, Goode ZH, Conaway JW, Conaway
            RC
  TITLE     Elongin functions as a loading factor for Mediator at 
            ATF6alpha-regulated ER stress response genes.
  JOURNAL   Proc Natl Acad Sci U S A. 2021 Sep 28;118(39):e2108751118. doi: 
            10.1073/pnas.2108751118.
  PUBMED    34544872
REFERENCE   2  (bases 1 to 6219)
  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"; journalName: "Proc Natl 
            Acad Sci U S A."; date: "2021-09-28"; volume: "118(39):e2108751118. 
            doi"; pages: " 10.1073/pnas.2108751118"
COMMENT     Sequence Label: pET-28a (+)-hTFIIB
FEATURES             Location/Qualifiers
     source          1..6219
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1124..1141)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             1160..1182
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1197..1221
                     /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(1221..1240)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1222..1240)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1324..1343)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1549..1626
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1627..2709
                     /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(1646..1665)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3518..3709
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3703..3725)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3811..3950
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3866..3885
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3965..3982)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4136..4724)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4216..4235)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4835..4854)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4846..5661
                     /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             4846..5661
                     /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(4919..4938)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(5753..6208)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5890..5911)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6102..6121
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagttatagc tgtggtagtt
      181 tgtccactgg ggtgtcaaat ttgaagtctg taggaaacag atctggggct cgaggataga
      241 tcagtctata ggactgtctg attgtaacat cagcaacacc agcaatatct ccaatttctt
      301 tttgggtcct cttttcagct gatgcctgtg aggccatgta aatagctgcc gctgccacag
      361 agatggggct cctcccagga accaagtcca attccacagc tttacgggct atatgtgtag
      421 ctgccatctg tacttgttta ggaagacaaa ggttggaaca gaacctggac atgaagtccc
      481 cagttgtaat caaatccaca ctggtttcta gcgctttcaa aataagttta aaacaccgac
      541 caatttcttt cttagaaatt cgtgatacgg cacatatttc tttaaatgtc ctaggaaccc
      601 cttcttgtct acaggcaata tagagacaag cagaagctat agcatcatta gctcttccct
      661 tcaggctctt ctgttcatat acttgcttga ataaattatt tgttcgatca actatatttc
      721 gaggtagatt gattctgtct gccatggtag tgatttcttt gaatgcattc atcattgccc
      781 gatcagaact gctcattgtt ctccgattct ggtacttaga attgccaaat tcgtcaaaac
      841 ttgcagctcc tgtgcccttg ccaatcatgg tagacaaatc tccatcactc agaagaggat
      901 tctgagaatc tccaactcga gatggatctt ttgttgcttt gtcattgctg aaagttcgcc
      961 attcagatcc cacatcaata acccggtcac ctacaaccaa gccacattca ggacagatca
     1021 tatcaccggc tctgtagtcc tccactaaaa tcgcatctgg atggtttgga catgtgactc
     1081 ttggaagagc atccaaacgg ctggtagacg ccatgccgct gctgtgatga tgatgatgat
     1141 ggctgctgcc catggtatat ctccttctta aagttaaaca aaattatttc tagaggggaa
     1201 ttgttatccg ctcacaattc ccctatagtg agtcgtatta atttcgcggg atcgagatct
     1261 cgatcctcta cgccggacgc atcgtggccg gcatcaccgg cgccacaggt gcggttgctg
     1321 gcgcctatat cgccgacatc accgatgggg aagatcgggc tcgccacttc gggctcatga
     1381 gcgcttgttt cggcgtgggt atggtggcag gccccgtggc cgggggactg ttgggcgcca
     1441 tctccttgca tgcaccattc cttgcggcgg cggtgctcaa cggcctcaac ctactactgg
     1501 gctgcttcct aatgcaggag tcgcataagg gagagcgtcg agatcccgga caccatcgaa
     1561 tggcgcaaaa cctttcgcgg tatggcatga tagcgcccgg aagagagtca attcagggtg
     1621 gtgaatgtga aaccagtaac gttatacgat gtcgcagagt atgccggtgt ctcttatcag
     1681 accgtttccc gcgtggtgaa ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg
     1741 gaagcggcga tggcggagct gaattacatt cccaaccgcg tggcacaaca actggcgggc
     1801 aaacagtcgt tgctgattgg cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa
     1861 attgtcgcgg cgattaaatc tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg
     1921 gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc
     1981 gtcagtgggc tgatcattaa ctatccgctg gatgaccagg atgccattgc tgtggaagct
     2041 gcctgcacta atgttccggc gttatttctt gatgtctctg accagacacc catcaacagt
     2101 attattttct cccatgaaga cggtacgcga ctgggcgtgg agcatctggt cgcattgggt
     2161 caccagcaaa tcgcgctgtt agcgggccca ttaagttctg tctcggcgcg tctgcgtctg
     2221 gctggctggc ataaatatct cactcgcaat caaattcagc cgatagcgga acgggaaggc
     2281 gactggagtg ccatgtccgg ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt
     2341 cccactgcga tgctggttgc caacgatcag atggcgctgg gcgcaatgcg cgccattacc
     2401 gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg gatacgacga taccgaagac
     2461 agctcatgtt atatcccgcc gttaaccacc atcaaacagg attttcgcct gctggggcaa
     2521 accagcgtgg accgcttgct gcaactctct cagggccagg cggtgaaggg caatcagctg
     2581 ttgcccgtct cactggtgaa aagaaaaacc accctggcgc ccaatacgca aaccgcctct
     2641 ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc
     2701 gggcagtgag cgcaacgcaa ttaatgtaag ttagctcact cattaggcac cgggatctcg
     2761 accgatgccc ttgagagcct tcaacccagt cagctccttc cggtgggcgc ggggcatgac
     2821 tatcgtcgcc gcacttatga ctgtcttctt tatcatgcaa ctcgtaggac aggtgccggc
     2881 agcgctctgg gtcattttcg gcgaggaccg ctttcgctgg agcgcgacga tgatcggcct
     2941 gtcgcttgcg gtattcggaa tcttgcacgc cctcgctcaa gccttcgtca ctggtcccgc
     3001 caccaaacgt ttcggcgaga agcaggccat tatcgccggc atggcggccc cacgggtgcg
     3061 catgatcgtg ctcctgtcgt tgaggacccg gctaggctgg cggggttgcc ttactggtta
     3121 gcagaatgaa tcaccgatac gcgagcgaac gtgaagcgac tgctgctgca aaacgtctgc
     3181 gacctgagca acaacatgaa tggtcttcgg tttccgtgtt tcgtaaagtc tggaaacgcg
     3241 gaagtcagcg ccctgcacca ttatgttccg gatctgcatc gcaggatgct gctggctacc
     3301 ctgtggaaca cctacatctg tattaacgaa gcgctggcat tgaccctgag tgatttttct
     3361 ctggtcccgc cgcatccata ccgccagttg tttaccctca caacgttcca gtaaccgggc
     3421 atgttcatca tcagtaaccc gtatcgtgag catcctctct cgtttcatcg gtatcattac
     3481 ccccatgaac agaaatcccc cttacacgga ggcatcagtg accaaacagg aaaaaaccgc
     3541 ccttaacatg gcccgcttta tcagaagcca gacattaacg cttctggaga aactcaacga
     3601 gctggacgcg gatgaacagg cagacatctg tgaatcgctt cacgaccacg ctgatgagct
     3661 ttaccgcagc tgcctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct
     3721 cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg
     3781 cgcgtcagcg ggtgttggcg ggtgtcgggg cgcagccatg acccagtcac gtagcgatag
     3841 cggagtgtat actggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat
     3901 tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg cgctcttccg
     3961 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     4021 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     4081 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     4141 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     4201 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     4261 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     4321 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     4381 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     4441 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     4501 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     4561 acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     4621 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     4681 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     4741 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     4801 acaataaaac tgtctgctta cataaacagt aatacaaggg gtgttatgag ccatattcaa
     4861 cgggaaacgt cttgctctag gccgcgatta aattccaaca tggatgctga tttatatggg
     4921 tataaatggg ctcgcgataa tgtcgggcaa tcaggtgcga caatctatcg attgtatggg
     4981 aagcccgatg cgccagagtt gtttctgaaa catggcaaag gtagcgttgc caatgatgtt
     5041 acagatgaga tggtcagact aaactggctg acggaattta tgcctcttcc gaccatcaag
     5101 cattttatcc gtactcctga tgatgcatgg ttactcacca ctgcgatccc cgggaaaaca
     5161 gcattccagg tattagaaga atatcctgat tcaggtgaaa atattgttga tgcgctggca
     5221 gtgttcctgc gccggttgca ttcgattcct gtttgtaatt gtccttttaa cagcgatcgc
     5281 gtatttcgtc tcgctcaggc gcaatcacga atgaataacg gtttggttga tgcgagtgat
     5341 tttgatgacg agcgtaatgg ctggcctgtt gaacaagtct ggaaagaaat gcataaactt
     5401 ttgccattct caccggattc agtcgtcact catggtgatt tctcacttga taaccttatt
     5461 tttgacgagg ggaaattaat aggttgtatt gatgttggac gagtcggaat cgcagaccga
     5521 taccaggatc ttgccatcct atggaactgc ctcggtgagt tttctccttc attacagaaa
     5581 cggctttttc aaaaatatgg tattgataat cctgatatga ataaattgca gtttcatttg
     5641 atgctcgatg agtttttcta agaattaatt catgagcgga tacatatttg aatgtattta
     5701 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctaaattgta
     5761 agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac
     5821 caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg
     5881 agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa
     5941 gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt
     6001 tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt
     6061 agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga
     6121 gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc
     6181 gcgcttaatg cgccgctaca gggcgcgtcc cattcgcca
//
