LOCUS       sequence-335551-        7647 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Co-expresses human TFIIF. The resulting plasmid was used to generate
            a expression construct encoding 2 subunits, with His-tag on RAP30. 
            Originally from MN Gonzalez, was submitted with permission.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7647)
  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 7647)
  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: pETDuet-TFIIF
FEATURES             Location/Qualifiers
     source          1..7647
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    3..27
                     /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             42..64
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             83..100
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        938..956
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    957..981
                     /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)."
     CDS             2596..2640
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     primer_bind     complement(2678..2696)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2692..2739
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2776..3231
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2863..2882)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3073..3094
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(3346..4137,4138..4206))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3969..3988
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4207..4299)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4380..4968
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4869..4888
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5122..5139
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5154..5293
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5219..5238)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5379..5401
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5395..5586)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(6158..7240)
                     /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     7202..7221
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7241..7318)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7524..7543
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        join(7631..7647,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct ctagaaataa ttttgtttaa ctttaagaag
       61 gagatatacc atgggcagca gccatcacca tcatcaccac agccaggatc cgaattcaat
      121 ggccgagcgc ggggaactcg acttgaccgg cgccaaacag aacacaggag tgtggctagt
      181 caaggttcct aaatatttgt cacagcaatg ggctaaagcc tctggaagag gtgaagttgg
      241 gaaactgcgg attgccaaga ctcaaggaag gactgaggtg tcatttactt tgaatgagga
      301 tcttgcaaat attcatgata ttggtggaaa accagcttca gtcagtgctc ctagagaaca
      361 tccatttgtc ttgcaaagtg ttggaggaca gacattaaca gtatttactg agagctcatc
      421 agataagctg tcattggaag gaatagtggt acaaagagct gaatgccgac cagctgccag
      481 tgaaaactac atgcgattaa aaagattgca aatagaagag tcttccaaac cagtgaggct
      541 atcacaacag ctggacaaag ttgtaacaac caattacaaa cctgttgcta atcatcaata
      601 caatatcgaa tatgaaagga aaaagaaaga agacggaaag cgagctcgag ctgataaaca
      661 acatgtttta gacatgctat tttcagcctt tgagaaacat caatactata atcttaagga
      721 cttggtggac atcacaaaac aacctgtggt gtacctgaag gaaatcttaa aagaaattgg
      781 tgttcagaat gtaaaaggga tccacaaaaa cacatgggag ctgaagccag agtacagaca
      841 ctatcaagga gaagaaaaga gtgactaagc ttgcggccgc ataatgctta agtcgaacag
      901 aaagtaatcg tattgtacac ggccgcataa tcgaaattaa tacgactcac tataggggaa
      961 ttgtgagcgg ataacaattc cccatcttag tatattagtt aagtataaga aggagatata
     1021 catatggcgg ccctaggccc tagcagccag aatgtcactg aatacgtcgt tcgagttcct
     1081 aagaatacaa ccaaaaaata taacatcatg gcttttaatg cagccgacaa agtcaacttt
     1141 gctacgtgga atcaggctcg gctggagcgg gacttgagca acaagaaaat ctaccaagag
     1201 gaggagatgc ccgaatcggg cgcgggcagt gagttcaacc gcaagcttcg ggaggaggct
     1261 cggaggaaga agtacggcat cgtcctcaag gagttccggc ccgaggacca gccctggctg
     1321 ctccgggtca acggcaaatc aggcaggaag ttcaagggca tcaagaaggg aggcgtaaca
     1381 gagaacacgt cctactacat cttcacccag tgccccgacg gggccttcga ggccttcccc
     1441 gtgcacaact ggtacaattt cacaccgctg gcccggcatc gcacgctcac tgccgaggag
     1501 gccgaggagg agtgggagag gaggaacaag gtgctgaacc acttcagcat catgcagcag
     1561 cggcggctca aggatcagga ccaggacgag gatgaggagg agaaggagaa acgtggccgc
     1621 aggaaggcga gcgagctgcg catccacgac ctggaggacg acctggagat gtcgtccgat
     1681 gccagtgatg ccagtggtga ggaggggggc agagtcccca aggccaagaa gaaggcgccg
     1741 ctggccaagg gcggcaggaa aaagaagaag aagaagggtt cagacgacga ggccttcgag
     1801 gacagcgatg atggggactt cgagggccaa gaggtggact acatgtcaga cggctccagt
     1861 agctcccaag aagagcctga gagcaaggcc aaggcgccgc agcaggagga ggggcccaag
     1921 ggtgtcgatg agcagagcga cagtagtgag gagagtgagg aggagaagcc gcctgaggag
     1981 gacaaggagg aggaggagga gaagaaggca cccaccccgc aggagaagaa gcgcaggaaa
     2041 gacagcagcg aggagtcgga cagctcagag gagagcgaca ttgacagcga ggcctcctca
     2101 gccctcttca tggcgaagaa gaagacgcca cccaagagag agcggaagcc gtcgggaggg
     2161 agctcaaggg gcaacagccg cccaggcacg cccagcgcag agggtggcag cacctcctcc
     2221 accctgcggg cggctgccag caaactcgag caagggaagc gggtgagcga gatgcctgca
     2281 gccaagcggt tgcggctgga cacgggaccc cagagcctgt ctgggaagtc gacaccccag
     2341 ccaccatcag gcaagacaac acccaacagc ggcgacgtgc aggtgactga ggatgccgtg
     2401 cgccgctacc tgacacggaa gcccatgacc actaaggacc tgctgaaaaa gttccagacc
     2461 aagaagacag ggctgagcag cgagcagaca gtgaacgtgt tggcccagat cctcaagcga
     2521 ctcaaccccg agcgcaagat gatcaacgac aaaatgcact tctccctcaa ggagtgaggt
     2581 accctcgagt ctggtaaaga aaccgctgct gcgaaatttg aacgccagca catggactcg
     2641 tctactagcg cagcttaatt aacctaggct gctgccaccg ctgagcaata actagcataa
     2701 ccccttgggg cctctaaacg ggtcttgagg ggttttttgc tgaaaggagg aactatatcc
     2761 ggattggcga atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     2821 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     2881 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     2941 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     3001 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     3061 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     3121 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     3181 tttaacaaaa atttaacgcg aattttaaca aaatattaac gtttacaatt tctggcggca
     3241 cgatggcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     3301 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     3361 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     3421 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     3481 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     3541 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     3601 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     3661 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     3721 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     3781 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     3841 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     3901 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     3961 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     4021 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     4081 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     4141 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     4201 actcatactc ttcctttttc aatcatgatt gaagcattta tcagggttat tgtctcatga
     4261 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggtcatgac caaaatccct
     4321 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     4381 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     4441 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     4501 agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg ccaccacttc
     4561 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     4621 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     4681 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     4741 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     4801 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     4861 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     4921 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     4981 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     5041 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     5101 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg
     5161 cggtattttc tccttacgca tctgtgcggt atttcacacc gcaatggtgc actctcagta
     5221 caatctgctc tgatgccgca tagttaagcc agtatacact ccgctatcgc tacgtgactg
     5281 ggtcatggct gcgccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
     5341 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
     5401 gttttcaccg tcatcaccga aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc
     5461 gtgaagcgat tcacagatgt ctgcctgttc atccgcgtcc agctcgttga gtttctccag
     5521 aagcgttaat gtctggcttc tgataaagcg ggccatgtta agggcggttt tttcctgttt
     5581 ggtcactgat gcctccgtgt aagggggatt tctgttcatg ggggtaatga taccgatgaa
     5641 acgagagagg atgctcacga tacgggttac tgatgatgaa catgcccggt tactggaacg
     5701 ttgtgagggt aaacaactgg cggtatggat gcggcgggac cagagaaaaa tcactcaggg
     5761 tcaatgccag cgcttcgtta atacagatgt aggtgttcca cagggtagcc agcagcatcc
     5821 tgcgatgcag atccggaaca taatggtgca gggcgctgac ttccgcgttt ccagacttta
     5881 cgaaacacgg aaaccgaaga ccattcatgt tgttgctcag gtcgcagacg ttttgcagca
     5941 gcagtcgctt cacgttcgct cgcgtatcgg tgattcattc tgctaaccag taaggcaacc
     6001 ccgccagcct agccgggtcc tcaacgacag gagcacgatc atgctagtca tgccccgcgc
     6061 ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgag atcccggtgc
     6121 ctaatgagtg agctaactta cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     6181 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     6241 tattgggcgc cagggtggtt tttcttttca ccagtgagac gggcaacagc tgattgccct
     6301 tcaccgcctg gccctgagag agttgcagca agcggtccac gctggtttgc cccagcaggc
     6361 gaaaatcctg tttgatggtg gttaacggcg ggatataaca tgagctgtct tcggtatcgt
     6421 cgtatcccac taccgagatg tccgcaccaa cgcgcagccc ggactcggta atggcgcgca
     6481 ttgcgcccag cgccatctga tcgttggcaa ccagcatcgc agtgggaacg atgccctcat
     6541 tcagcatttg catggtttgt tgaaaaccgg acatggcact ccagtcgcct tcccgttccg
     6601 ctatcggctg aatttgattg cgagtgagat atttatgcca gccagccaga cgcagacgcg
     6661 ccgagacaga acttaatggg cccgctaaca gcgcgatttg ctggtgaccc aatgcgacca
     6721 gatgctccac gcccagtcgc gtaccgtctt catgggagaa aataatactg ttgatgggtg
     6781 tctggtcaga gacatcaaga aataacgccg gaacattagt gcaggcagct tccacagcaa
     6841 tggcatcctg gtcatccagc ggatagttaa tgatcagccc actgacgcgt tgcgcgagaa
     6901 gattgtgcac cgccgcttta caggcttcga cgccgcttcg ttctaccatc gacaccacca
     6961 cgctggcacc cagttgatcg gcgcgagatt taatcgccgc gacaatttgc gacggcgcgt
     7021 gcagggccag actggaggtg gcaacgccaa tcagcaacga ctgtttgccc gccagttgtt
     7081 gtgccacgcg gttgggaatg taattcagct ccgccatcgc cgcttccact ttttcccgcg
     7141 ttttcgcaga aacgtggctg gcctggttca ccacgcggga aacggtctga taagagacac
     7201 cggcatactc tgcgacatcg tataacgtta ctggtttcac attcaccacc ctgaattgac
     7261 tctcttccgg gcgctatcat gccataccgc gaaaggtttt gcgccattcg atggtgtccg
     7321 ggatctcgac gctctccctt atgcgactcc tgcattagga agcagcccag tagtaggttg
     7381 aggccgttga gcaccgccgc cgcaaggaat ggtgcatgca aggagatggc gcccaacagt
     7441 cccccggcca cggggcctgc caccataccc acgccgaaac aagcgctcat gagcccgaag
     7501 tggcgagccc gatcttcccc atcggtgatg tcggcgatat aggcgccagc aaccgcacct
     7561 gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga ggatcgagat cgatctcgat
     7621 cccgcgaaat taatacgact cactata
//
