LOCUS       sequence-421746-        5716 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TEFM protein expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5716)
  AUTHORS   Sultana S, Solotchi M, Ramachandran A, Patel SS
  TITLE     Transcriptional fidelities of human mitochondrial POLRMT, yeast 
            mitochondrial Rpo41, and phage T7 single-subunit RNA polymerases.
  JOURNAL   J Biol Chem. 2017 Nov 3;292(44):18145-18160. doi: 
            10.1074/jbc.M117.797480. Epub 2017 Sep 7.
  PUBMED    28882896
REFERENCE   2  (bases 1 to 5716)
  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: 
            "10.1074/jbc.M117.797480"; journalName: "J Biol Chem"; date: 
            "2017-11-3- 3"; volume: "292"; issue: "44"; pages: "18145-18160"
COMMENT     Sequence Label: mt TEFM
FEATURES             Location/Qualifiers
     source          1..5716
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     547..566
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(586..663)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     primer_bind     complement(741..763)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     936..955
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      993..1421
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1080..1099)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1290..1311
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1498..1515)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1669..2257)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1749..1768)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2428..3219,3220..3288))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     3051..3070
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3289..3380)
                     /gene="bla"
                     /label=AmpR promoter
     terminator      complement(3399..3426)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3518..3604)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     3757..3774
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     CDS             complement(4842..4862)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             complement(4884..4901)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4923..4945)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4934..4950
                     /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             complement(join(5219..5716,1..585))
                     /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 acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt gacccaatgc
       61 gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa tactgttgat
      121 gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg cagcttccac
      181 agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga cgcgttgcgc
      241 gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta ccatcgacac
      301 caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa tttgcgacgg
      361 cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt tgcccgccag
      421 ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt ccactttttc
      481 ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg tctgataaga
      541 gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca ccaccctgaa
      601 ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcacc attcgatggt
      661 gtcaacgtaa atgcatgccg cttcgccttc gcgcgcgaat tgatctgctg cctcgcgcgt
      721 ttcggtgatg acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt
      781 ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg
      841 tgtcggggcg cagccatgac ccagtcacgt agcgatagcg gagtgtacca gtcacgtagc
      901 gatatcgcta cgtgactggt actggcttaa ctatgcggca tcagagcaga ttgtactgag
      961 agtgcaccat agcagcctga atggcgaatg ggacgcgccc tgtagcggcg cattaagcgc
     1021 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     1081 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     1141 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     1201 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     1261 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     1321 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     1381 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa ttatgcggtg
     1441 tgaaataccg cacagatgcg taaggagaaa ataccgcatc aggcgctctt ccgcttcctc
     1501 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     1561 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     1621 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     1681 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     1741 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     1801 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     1861 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     1921 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     1981 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     2041 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     2101 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     2161 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     2221 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     2281 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     2341 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     2401 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     2461 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
     2521 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     2581 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     2641 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     2701 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     2761 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     2821 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     2881 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     2941 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     3001 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaacacggg ataataccgc
     3061 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     3121 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     3181 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     3241 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     3301 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     3361 tatttagaaa aataaacaaa agagtttgta gaaacgcaaa aaggccatcc gtcaggatgg
     3421 ccttctgctt aatttgatgc ctggcagttt atggcgggcg tcctgcccgc caccctccgg
     3481 gccgttgctt cgcaacgttc aaatccgctc ccggcggatt tgtcctactc aggagagcgt
     3541 tcaccgacaa acaacagata aaacgaaagg cccagtcttt cgactgagcc tttcgtttta
     3601 tttgatgcct ggcagttccc tactctcgca tggggagacc ccacactacc atcggcgcta
     3661 cggcgtttca cttctgagtt cggcatgggg tcaggtggga ccaccgcgct actgccgcca
     3721 ggcaaattct gttttatcag accgcttctg cgttctgatt taatctgtat caggctgaaa
     3781 atcttctctc atccgccaaa acagccaagc ttggtaccgc atgcctcgag actgcaggct
     3841 ctagattcga aagcggccgc tcacggttgg gagtcaaaaa cagccagttc ataaaatgcg
     3901 atagcttgca gcagcgagtc gtacagttct tcaacgcgtt gcagttcggt ggacaggaac
     3961 atctgacgat agtgaacgat tttatcactc gggaaaaaca cacgcgggtc cgctttcaga
     4021 atagaatcga acagaaactg tttcaccagt tctttaccgc tggtgcgcga gtcgccgatc
     4081 atcagttcga agtgtttacc gacagcatta cggttcatgc tcagaacttg atgctggcca
     4141 tcctgtgcaa acgttttgtt cagcagagcg tacagcattg cttccatgat gtggaaatgc
     4201 agcagaatcg gaaacagact agaattctga atcgacagac cggttttttc caggacataa
     4261 aaatcggctt tcggcatttt tgagataatg ctcgagattt cttccaggta aactgaggaa
     4321 ctataaatac cgcgcatcag actccaacgg tcagattgct gccaatccag aaccgtcagt
     4381 ttgcggtcca gatgtgccca ggcaatgcga cgggtaccaa aaacaattga gataatggaa
     4441 ttgaccgctt tcaggcgttc acgttcgata tccggtttca gcagtttgcg cagaaaacgg
     4501 ttttccggac ttttgcgttt ttcacggccc gttttcggac acagaattga attgcaaact
     4561 tgcacggtgg atttatattt aaacagcggc acgttcatca gagattccag attctggaac
     4621 ggaccaaagt tttcacggtg ttcgacgatg ttaattgagc gacggccacg cagcagacgg
     4681 aatgcttcca gttctttcgt gctcgcggta ttcagcacat gcagaatcga tgcttgctgt
     4741 tcagagctaa acagtttatc cagggcattt tccggttctt tagcgttttc gtcgcagaag
     4801 gtaacattcg gggtgatttt cttcggggtg gttgatttgg cgccctgaaa atacaggttt
     4861 tcggtcgttg ggatatcgta atcgtgatgg tgatggtgat ggtagtacga catggtctgt
     4921 ttcctgtgtg aaattgttat ccgctcacaa ttccacagat tatacggacc ggatgattaa
     4981 ttgtcaacag ctcatttcag aatatttgcc agaaccgtta tgatgtcggc gcaaaaaaca
     5041 ttatccagaa cgggagtgcg ccttgagcga cacgaattat gcagtgattt acgacctgca
     5101 cagccatacc acagcttccg atggctgcct gacgccagaa gcattggtgc accgtgcagt
     5161 cgatgataag ctgtcaaacc agatcaattc gcgctaactt acattaattg cgttgcgctc
     5221 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     5281 cgcggggaga ggcggtttgc gtattgggtg ccagggtggt ttttcttttc accagtgaga
     5341 cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca
     5401 cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac
     5461 atgagctgtc ttcggtatcg tcgtatccca ctaccgagat atccgcacca acgcgcagcc
     5521 cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg
     5581 cagtgggaac gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac
     5641 tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc
     5701 agccagccag acgcag
//
