LOCUS       sequence-429704-        6704 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses his-tagged Mut5 reverse transcriptase in E. coli hosts.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6704)
  TITLE     Bacterial expression plasmids for production of recombinant PNK
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6704)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pET-hisM5RT
FEATURES             Location/Qualifiers
     source          1..6704
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..15,16..32,33..38)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     terminator      complement(404..451)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     447..465
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(2589..2606)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             2626..2648
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     primer_bind     complement(2679..2698)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2680..2698)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2782..2801)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             4259..4450
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(4444..4466)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    4552..4691
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4607..4626
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4706..4723)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4877..5465)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4957..4976)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5636..6427,6428..6496))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6259..6278
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6497..6601)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6669..6687
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ttctcatgtt tgacagctta tcatcgataa gctttaatgc ggtagtttat cacagttaaa
       61 ttgctaacgc agtcaggcac cgtgtatgaa atctaacaat gcgctcatcg tcatcctcgg
      121 caccgtcacc ctggatgctg taggcatagg cttggttatg ccggtactgc cgggcctctt
      181 gcgggatatc gtccattccg acagcatcgc cagtcactat ggcgtgctgc tagcgctata
      241 tgcgttgatg caatttctat gcgcacccgt tctcggagca ctgtccgacc gctttggccg
      301 ccgcccagtc ctgctcgctt cgctacttgg agccactatc gactacgcga tcatggcgac
      361 cacacccgtc ctgtggatat ccggatatag ttcctccttt cagcaaaaaa cccctcaaga
      421 cccgtttaga ggccccaagg ggttatgcta gttattgctc agcggtggca gcagccaact
      481 cagcttcctt tcgggctttg ttagcagccg gatccttaat taattaagcg ggaattgggt
      541 gatgaatttt ctatgaggag ggtagaggtg tctggagtct ctgtgatggc tgcctttcgg
      601 gccgcttggt cagccatccg gttgcctcta gcctcggcgc tgtgtccctt ttgatgtcct
      661 ggacaatgga ttatgctaag tcttttgggc agaaagaggg cttttagtag ggccaagatc
      721 tcgtctttat ttttgatctc tttgcctttt gatgtgagcc acccacgcct tctgtatatt
      781 tctccatgga tatgggcagt agcaaaagca taacggctat tagtataaac atttagcttc
      841 ttaccttctg ccatctttag ggcctgggtg agtgctatca gttcagcccg ctgagcggat
      901 gtcccggctg gcagggcttt agcccagatt acctcggtct cggtggtcac cgcagctccc
      961 gccttacgct gtccctcttg taagagactg cttccacccg tgtaccaggt gtggtcggcg
     1021 tctgggagcg gctggtccgt taggtcgggt cgggttccgt gggcttcggc caggatatca
     1081 aggcagttgt gttgcagccc ttcctcaggc agtgggagca gcgtagccgg gttcagggct
     1141 accaccggtc cgaactggac ccggtccgtg tccaaaagca aggcctgata gtgagtcatc
     1201 cgggcgttgg aaagccagcg gtcggggggt tgtttgacta gtgcctctac tgcatggggg
     1261 gccagaatga ctagtggctg tcccatggtt agcttgcctg catcctttgt cagtacggca
     1321 atggctgcta ccatccgtag gcaagggggc cacccagctg ctactgggtc tagctttttg
     1381 gacaggtagg ccaccggccg acgccaaggt cccagttttt gcgttaggac acctttggcg
     1441 tagccctgct tctcgtcgac aaagagttca aagggcttag tcaaatctgg caaccccagg
     1501 gctggggcag ttagaagagc ttgcttgatt tcttgatagg ccttttgttg gtctgggccc
     1561 caattaaaca gagtccccgg tttggtgaga gggtacaagg gggctgccat ttctgcaaac
     1621 ccagggatcc agaggcgaca gaagcctgcc gtccctagga actcccttag ttgtcgaggg
     1681 gtcttcggag taggctgccc catcacagtc tcttttctgg cctcagtcag ccatctatga
     1741 ccctctttta gaagataccc cagatacttg acctgtttct ggcaaatttg ggctttcttg
     1801 gccgaggccc gatacccgag gttccctagg gtttgtaaca gggcccgagt accttgttgg
     1861 cagtctagct cagaagtggc ggccagcagt aagtcatcca cgtactgtag caggatcaag
     1921 tctgggtgct ggatccggaa gtctgctagg tctctgtgca gtgcctcatt aaacagggtg
     1981 ggactgtttt tgaaaccctg tgggagtctg gtccaggtca attgtcctga gattcccatc
     2041 tctggatctc tccactcaaa ggcgaagaga ggctgactgg tggggtggag tctcaggcag
     2101 aaaaaggcat cctttaaatc aagcacagtg taccactggt gggacggtgg gggcccgctc
     2161 aagaggttgt aagggttggg cacggtgggg tggatgtctt ccacccgctt gttgacttct
     2221 ctcagatcct ggacaggcct ataatcatta gtccctggtt tcttaacggg tagcaggggc
     2281 gtgttccagg gggactggca gggtaccagt attccctggt ccaacagtct ctgtatgtgg
     2341 ggcttgatcc ccagtctggc ttcttgtgac atggggtatt gttttatgga cacgggggta
     2401 gaggttgctt tcagaggtat gatcagagga gcttggcgaa ctgccagtcc catgcccccg
     2461 gtttccgccc aggcctgagg aaaatcagac agccatgtgg accctagaga aacatctggc
     2521 tcttttgagg tctcatgtag ccgatgctca tcttctatat ttagggtcat gcccatggaa
     2581 ccagatccat gatgatggtg gtggtgcgat cctctcatat gtatatctcc ttcttaaagt
     2641 taaacaaaat tatttctaga gggaaaccgt tgtggtctcc ctatagtgag tcgtattaat
     2701 ttcgcgggat cgagatctcg atcctctacg ccggacgcat cgtggccggc atcaccggcg
     2761 ccacaggtgc ggttgctggc gcctatatcg ccgacatcac cgatggggaa gatcgggctc
     2821 gccacttcgg gctcatgagc gcttgtttcg gcgtgggtat ggtggcaggc cccgtggccg
     2881 ggggactgtt gggcgccatc tccttgcatg caccattcct tgcggcggcg gtgctcaacg
     2941 gcctcaacct actactgggc tgcttcctaa tgcaggagtc gcataaggga gagcgtcgac
     3001 cgatgccctt gagagccttc aacccagtca gctccttccg gtgggcgcgg ggcatgacta
     3061 tcgtcgccgc acttatgact gtcttcttta tcatgcaact cgtaggacag gtgccggcag
     3121 cgctctgggt cattttcggc gaggaccgct ttcgctggag cgcgacgatg atcggcctgt
     3181 cgcttgcggt attcggaatc ttgcacgccc tcgctcaagc cttcgtcact ggtcccgcca
     3241 ccaaacgttt cggcgagaag caggccatta tcgccggcat ggcggccgac gcgctgggct
     3301 acgtcttgct ggcgttcgcg acgcgaggct ggatggcctt ccccattatg attcttctcg
     3361 cttccggcgg catcgggatg cccgcgttgc aggccatgct gtccaggcag gtagatgacg
     3421 accatcaggg acagcttcaa ggatcgctcg cggctcttac cagcctaact tcgatcattg
     3481 gaccgctgat cgtcacggcg atttatgccg cctcggcgag cacatggaac gggttggcat
     3541 ggattgtagg cgccgcccta taccttgtct gcctccccgc gttgcgtcgc ggtgcatgga
     3601 gccgggccac ctcgacctga atggaagccg gcggcacctc gctaacggat tcaccactcc
     3661 aagaattgga gccaatcaat tcttgcggag aactgtgaat gcgcaaacca acccttggca
     3721 gaacatatcc atcgcgtccg ccatctccag cagccgcacg cggcgcatct cgggcagcgt
     3781 tgggtcctgg ccacgggtgc gcatgatcgt gctcctgtcg ttgaggaccc ggctaggctg
     3841 gcggggttgc cttactggtt agcagaatga atcaccgata cgcgagcgaa cgtgaagcga
     3901 ctgctgctgc aaaacgtctg cgacctgagc aacaacatga atggtcttcg gtttccgtgt
     3961 ttcgtaaagt ctggaaacgc ggaagtcagc gccctgcacc attatgttcc ggatctgcat
     4021 cgcaggatgc tgctggctac cctgtggaac acctacatct gtattaacga agcgctggca
     4081 ttgaccctga gtgatttttc tctggtcccg ccgcatccat accgccagtt gtttaccctc
     4141 acaacgttcc agtaaccggg catgttcatc atcagtaacc cgtatcgtga gcatcctctc
     4201 tcgtttcatc ggtatcatta cccccatgaa cagaaatccc ccttacacgg aggcatcagt
     4261 gaccaaacag gaaaaaaccg cccttaacat ggcccgcttt atcagaagcc agacattaac
     4321 gcttctggag aaactcaacg agctggacgc ggatgaacag gcagacatct gtgaatcgct
     4381 tcacgaccac gctgatgagc tttaccgcag ctgcctcgcg cgtttcggtg atgacggtga
     4441 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
     4501 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gcgcagccat
     4561 gacccagtca cgtagcgata gcggagtgta tactggctta actatgcggc atcagagcag
     4621 attgtactga gagtgcacca ttgcggtgtg aaataccgca cagatgcgta aggagaaaat
     4681 accgcatcag gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     4741 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     4801 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     4861 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     4921 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     4981 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     5041 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     5101 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     5161 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     5221 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     5281 tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc
     5341 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     5401 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     5461 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     5521 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     5581 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     5641 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     5701 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     5761 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     5821 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     5881 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     5941 ttgccattgc tgcaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     6001 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     6061 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     6121 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     6181 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     6241 gcccggcgtc aacacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     6301 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     6361 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     6421 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     6481 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     6541 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     6601 gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa
     6661 cctataaaaa taggcgtatc acgaggccct ttcgtcttca agaa
//
