LOCUS       sequence-413717-        7592 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TFB2M protein expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7592)
  AUTHORS   Basu U, Lee SW, Deshpande A, Shen J, Sohn BK, Cho H, Kim H, Patel SS
  TITLE     The C-terminal tail of the yeast mitochondrial transcription factor 
            Mtf1 coordinates template strand alignment, DNA scrunching and 
            timely transition into elongation.
  JOURNAL   Nucleic Acids Res. 2020 Mar 18;48(5):2604-2620. doi: 
            10.1093/nar/gkaa040.
  PUBMED    31980825
REFERENCE   2  (bases 1 to 7592)
  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"; doi: "10"; journalName: 
            "Nucleic Acids Res"; date: "2020-03-18- 18"; volume: "48"; issue: 
            "5"; pages: "2604-2620"
COMMENT     Sequence Label: mt deltaN20 TFB2MdeltaC7
FEATURES             Location/Qualifiers
     source          1..7592
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     108..127
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(147..224)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     430..449
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     533..552
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        533..551
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    552..576
                     /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             591..613
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             632..649
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             659..676
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             680..1780
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELVKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     1754..1777
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1832..1852
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2990..3007
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3060..3078)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3074..3121
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      3158..3613
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3245..3264)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3455..3476
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3705..4520)
                     /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(3705..4520)
                     /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     4428..4447
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4512..4531
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4642..5229
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5130..5149
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5383..5400
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5415..5554
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5480..5499)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5640..5662
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5656..5847)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(join(6656..7592,1..146))
                     /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"
ORIGIN
        1 ggaatgtaat tcagctccgc catcgccgct tccacttttt cccgcgtttt cgcagaaacg
       61 tggctggcct ggttcaccac gcgggaaacg gtctgataag agacaccggc atactctgcg
      121 acatcgtata acgttactgg tttcacattc accaccctga attgactctc ttccgggcgc
      181 tatcatgcca taccgcgaaa ggttttgcgc cattcgatgg tgtccgggat ctcgacgctc
      241 tcccttatgc gactcctgca ttaggaagca gcccagtagt aggttgaggc cgttgagcac
      301 cgccgccgca aggaatggtg catgcaagga gatggcgccc aacagtcccc cggccacggg
      361 gcctgccacc atacccacgc cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc
      421 ttccccatcg gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg cgccggtgat
      481 gccggccacg atgcgtccgg cgtagaggat cgagatctcg atcccgcgaa attaatacga
      541 ctcactatag gggaattgtg agcggataac aattcccctc tagaaataat tttgtttaac
      601 tttaagaagg agatatacca tgggcagcag ccatcatcat catcatcaca gcagcggcct
      661 ggtgccgcgc ggcagccaca tgaaaatcga agaaggtaaa ctggtaatct ggattaacgg
      721 cgataaaggc tataacggtc tcgctgaagt cggtaagaaa ttcgagaaag ataccggaat
      781 taaagtcacc gttgagcatc cggataaact ggaagagaaa ttcccacagg ttgcggcaac
      841 tggcgatggc cctgacatta tcttctgggc acacgaccgc tttggtggct acgctcaatc
      901 tggcctgttg gctgaaatca ccccggacaa agcgttccag gacaagctgt atccgtttac
      961 ctgggatgcc gtacgttaca acggcaagct gattgcttac ccgatcgctg ttgaagcgtt
     1021 atcgctgatt tataacaaag atctgctgcc gaacccgcca aaaacctggg aagagatccc
     1081 ggcgctggat aaagaactga aagcgaaagg taagagcgcg ctgatgttca acctgcaaga
     1141 accgtacttc acctggccgc tgattgctgc tgacgggggt tatgcgttca agtatgaaaa
     1201 cggcaagtac gacattaaag acgtgggcgt ggataacgct ggcgcgaaag cgggtctgac
     1261 cttcctggtt gacctgatta aaaacaaaca catgaatgca gacaccgatt actccatcgc
     1321 agaagctgcc tttaataaag gcgaaacagc gatgaccatc aacggcccgt gggcatggtc
     1381 caacatcgac accagcaaag tgaattatgg tgtaacggta ctgccgacct tcaagggtca
     1441 accatccaaa ccgttcgttg gtgtgctgag cgcaggtatt aacgccgcca gtccgaacaa
     1501 agagctggca aaagagttcc tcgaaaacta tctgctgact gatgaaggtc tggaagcggt
     1561 taataaagac aaaccgctgg gtgccgtagc gctgaagtct tacgaggaag agttggtgaa
     1621 agatccgcgg attgccgcca ctatggaaaa cgcccagaaa ggtgaaatca tgccgaacat
     1681 cccgcagatg tccgctttct ggtatgccgt gcgtactgcg gtgatcaacg ccgccagcgg
     1741 tcgtcagact gtcgatgaag ccctgaaaga cgcgcagact aattcgagct cgaacaacaa
     1801 caacaataac aataacaaca acctcgggga tgaaaactta tatttccaag gtgctagcgg
     1861 tcgcttttgc attttagggt ctgaagcggc gacgcgaaag catttgccgg cgaggaacca
     1921 ctgtgggctc tctgactcct ctccgcagct gtggcccgaa ccggatttca ggaatccgcc
     1981 aaggaaggcg tctaaggcca gcttagactt taagcgttac gtaaccgatc ggagattggc
     2041 tgagaccctg gcgcaaatct atttgggaaa accaagtaga cctccacacc tactgctgga
     2101 gtgcaatcca ggtcctggaa tcctgactca ggcattactt gaagctggtg ccaaagtggt
     2161 tgcgctcgaa agtgacaaaa cttttattcc acatttggag tccttaggaa aaaatctgga
     2221 tggaaaacta cgagtgattc actgtgactt ctttaaacta gatcctagaa gtggtggagt
     2281 aataaaacca cctgctatgt cttctcgagg gctctttaag aatttgggaa tagaagcagt
     2341 tccttggaca gcagacatcc ctttaaaagt agttggaatg ttcccaagta gaggtgagaa
     2401 aagggcactt tggaaactcg catatgactt gtattcctgt acttctatat ataaatttgg
     2461 acgaatagaa gtaaatatgt ttattggtga aaaagaattc cagaaactaa tggcagatcc
     2521 tggaaatcca gacttgtatc atgtattaag tgttatctgg caattagctt gtgagattaa
     2581 ggttctgcac atggagcctt ggtcatcatt tgatatatac acccggaaag ggccgctgga
     2641 aaacccaaag cgtagggaat tattagacca attacaacaa aagctgtatc ttattcaaat
     2701 gattcctcgt caaaatttat ttaccaagaa cttaacacct atgaactata atatattttt
     2761 tcacttgtta aagcactgtt ttgggaggcg cagcgccact gtaatagacc acttacgttc
     2821 attgactcca cttgatgcga gagatatatt gatgcaaata ggaaaacagg aggatgagaa
     2881 agtagttaac atgcaccctc aagacttcaa aacacttttt gaaactatag agcgttccaa
     2941 agattgtgct tataaatggc tgtattagaa gcttgcggcc gcactcgagc accaccacca
     3001 ccaccactga gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg ctgctgccac
     3061 cgctgagcaa taactagcat aaccccttgg ggcctctaaa cgggtcttga ggggtttttt
     3121 gctgaaagga ggaactatat ccggattggc gaatgggacg cgccctgtag cggcgcatta
     3181 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg
     3241 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     3301 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     3361 aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     3421 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     3481 acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc
     3541 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta
     3601 acgcttacaa tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     3661 ttttctaaat acattcaaat atgtatccgc tcatgaatta attcttagaa aaactcatcg
     3721 agcatcaaat gaaactgcaa tttattcata tcaggattat caataccata tttttgaaaa
     3781 agccgtttct gtaatgaagg agaaaactca ccgaggcagt tccataggat ggcaagatcc
     3841 tggtatcggt ctgcgattcc gactcgtcca acatcaatac aacctattaa tttcccctcg
     3901 tcaaaaataa ggttatcaag tgagaaatca ccatgagtga cgactgaatc cggtgagaat
     3961 ggcaaaagtt tatgcatttc tttccagact tgttcaacag gccagccatt acgctcgtca
     4021 tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg attgcgcctg agcgagacga
     4081 aatacgcgat cgctgttaaa aggacaatta caaacaggaa tcgaatgcaa ccggcgcagg
     4141 aacactgcca gcgcatcaac aatattttca cctgaatcag gatattcttc taatacctgg
     4201 aatgctgttt tcccggggat cgcagtggtg agtaaccatg catcatcagg agtacggata
     4261 aaatgcttga tggtcggaag aggcataaat tccgtcagcc agtttagtct gaccatctca
     4321 tctgtaacat cattggcaac gctacctttg ccatgtttca gaaacaactc tggcgcatcg
     4381 ggcttcccat acaatcgata gattgtcgca cctgattgcc cgacattatc gcgagcccat
     4441 ttatacccat ataaatcagc atccatgttg gaatttaatc gcggcctaga gcaagacgtt
     4501 tcccgttgaa tatggctcat aacacccctt gtattactgt ttatgtaagc agacagtttt
     4561 attgttcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     4621 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     4681 aacaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     4741 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag
     4801 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     4861 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     4921 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     4981 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     5041 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     5101 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     5161 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     5221 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     5281 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
     5341 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
     5401 gaagcggaag agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     5461 cgcaatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac
     5521 tccgctatcg ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga
     5581 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     5641 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagctgcg
     5701 gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt catccgcgtc
     5761 cagctcgttg agtttctcca gaagcgttaa tgtctggctt ctgataaagc gggccatgtt
     5821 aagggcggtt ttttcctgtt tggtcactga tgcctccgtg taagggggat ttctgttcat
     5881 gggggtaatg ataccgatga aacgagagag gatgctcacg atacgggtta ctgatgatga
     5941 acatgcccgg ttactggaac gttgtgaggg taaacaactg gcggtatgga tgcggcggga
     6001 ccagagaaaa atcactcagg gtcaatgcca gcgcttcgtt aatacagatg taggtgttcc
     6061 acagggtagc cagcagcatc ctgcgatgca gatccggaac ataatggtgc agggcgctga
     6121 cttccgcgtt tccagacttt acgaaacacg gaaaccgaag accattcatg ttgttgctca
     6181 ggtcgcagac gttttgcagc agcagtcgct tcacgttcgc tcgcgtatcg gtgattcatt
     6241 ctgctaacca gtaaggcaac cccgccagcc tagccgggtc ctcaacgaca ggagcacgat
     6301 catgcgcacc cgtggggccg ccatgccggc gataatggcc tgcttctcgc cgaaacgttt
     6361 ggtggcggga ccagtgacga aggcttgagc gagggcgtgc aagattccga ataccgcaag
     6421 cgacaggccg atcatcgtcg cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag
     6481 cgctgccggc acctgtccta cgagttgcat gataaagaag acagtcataa gtgcggcgac
     6541 gatagtcatg ccccgcgccc accggaagga gctgactggg ttgaaggctc tcaagggcat
     6601 cggtcgagat cccggtgcct aatgagtgag ctaacttaca ttaattgcgt tgcgctcact
     6661 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     6721 ggggagaggc ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg
     6781 gcaacagctg attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc
     6841 tggtttgccc cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg
     6901 agctgtcttc ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg cgcagcccgg
     6961 actcggtaat ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag
     7021 tgggaacgat gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc
     7081 agtcgccttc ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc
     7141 cagccagacg cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct
     7201 ggtgacccaa tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa
     7261 taatactgtt gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc
     7321 aggcagcttc cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac
     7381 tgacgcgttg cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt
     7441 ctaccatcga caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga
     7501 caatttgcga cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact
     7561 gtttgcccgc cagttgttgt gccacgcggt tg
//
