LOCUS       sequence-298742-        7789 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of MMLV reverse transcriptase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7789)
  TITLE     COVID-19 Diagnostic Toolkit Enzymes
REFERENCE   2  (bases 1 to 7789)
  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"
COMMENT     Sequence Label: pET-19b_MMLV-RT
FEATURES             Location/Qualifiers
     source          1..7789
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     101..120
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     204..223
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        204..222
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    223..247
                     /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             262..284
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             297..326
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             342..356
                     /codon_start=1
                     /product="enterokinase recognition and cleavage site"
                     /label=enterokinase site
                     /translation="DDDDK"
     primer_bind     complement(2493..2511)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2507..2554
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(2729..2734,2735..2751,2752..2757))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(2784..2802)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2870..2974
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2975..3043,3044..3835)
                     /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     complement(3193..3212)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4006..4594
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4495..4514
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4748..4765
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4780..4919
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4845..4864)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5005..5027
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5021..5212)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(6524..7606)
                     /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     7568..7587
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7607..7684)
                     /gene="lacI"
                     /label=lacI promoter
ORIGIN
        1 agatggcgcc caacagtccc ccggccacgg ggcctgccac catacccacg ccgaaacaag
       61 cgctcatgag cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg gcgatatagg
      121 cgccagcaac cgcacctgtg gcgccggtga tgccggccac gatgcgtccg gcgtagagga
      181 tcgagatctc gatcccgcga aattaatacg actcactata ggggaattgt gagcggataa
      241 caattcccct ctagaaataa ttttgtttaa ctttaagaag gagatatacc atgggccatc
      301 atcatcatca tcatcatcat catcacagca gcggccatat cgacgacgac gacaagcata
      361 tgggccagcc gctccaagtg ctcaccctga acatcgaaga cgaacatcgc ttgcacgaaa
      421 ccagcaaaga accggatgtt tcactgggtt ccacttggct gtcggacttt ccgcaagcgt
      481 gggcagaaac cggcggtatg ggtctggctg tgcgccaagc tccgctgatt atcccactga
      541 aggcaactag taccccggta tccattaaac agtacccgat gagccaggag gcccgtctgg
      601 gcattaaacc gcatatccag cgtctgctgg accagggtat cttagtcccg tgccagtccc
      661 cgtggaacac cccgctgctg ccggttaaaa aaccgggcac caacgactat cgcccggtac
      721 aagatttacg tgaagttaac aagcgcgtag aagatatcca tccgaccgtt cccaacccgt
      781 acaatctgct ctcaggtccg ccgccttccc accaatggta caccgttctg gacctgaaag
      841 atgccttctt ttgcttgcgt ctgcatccga ctagccagcc cctgtttgca ttcgagtggc
      901 gtgacccgga aatggggatc agcggccagc tcacctggac ccgtctgccg caaggtttca
      961 aaaactcacc taccctgttt aatgaagctc tgcaccgcga tctggcggat ttccgcattc
     1021 aacacccgga tctgattctg ctgcagtatg tggacgacct gctgctggcg gctacctccg
     1081 aactggattg ccagcagggc acccgtgcct tactgcaaac cctgggtaac ttaggttatc
     1141 gtgctagcgc gaaaaaagcc cagatctgtc agaagcaggt taaatacctg ggttacctcc
     1201 tgaaagaagg ccagcgctgg ttaaccgaag ctcgcaagga gacggtcatg ggtcagccaa
     1261 ccccgaaaac cccgcgccag ctgcgtgagt tcctgggcac cgcgggcttt tgtcgcctgt
     1321 ggatcccggg cttcgcggaa atggctgcgc cgctgtaccc gctcaccaaa ccggggaccc
     1381 tgttcaactg gggtccggat cagcaaaagg cttaccagga gatcaaacag gcgttgctga
     1441 ccgcaccagc tctcggtctg ccggacctga cgaaaccgtt tgaactgttt gtggacgaaa
     1501 aacagggtta cgcaaaaggt gttctgaccc agaaactggg tccatggcgt cgcccggtcg
     1561 cctacctgag caagaaactg gacccggttg cagctggctg gccgccgtgt ctccgtatgg
     1621 tagctgcaat cgctgtgctg accaaagatg cgggaaaact gaccatgggt caaccgttgg
     1681 tgatcctggc accgcatgcg gtagaagccc tggtgaaaca gccgccggac cgttggctgt
     1741 ccaacgcccg catgacgcac taccaggcgc tgctgctgga taccgatcgt gttcagttcg
     1801 gcccggttgt tgcgctgaac ccggcaactc tgcttccgtt accggaagag ggtctgcagc
     1861 acaactgcct tgatatcctc gcagaagcgc acggcacccg tcctgacctg acggaccagc
     1921 cactgccgga cgcagatcat acctggtaca ccgacggcag cagcctgctg caagagggcc
     1981 aacgcaaagc gggggcagcc gtaaccactg agaccgaggt tatttgggct aaagctctgc
     2041 cagccggcac ttctgcacag cgtgcggaac tgatcgctct gacccaagcc ctgaaaatgg
     2101 ctgaaggcaa gaaattgaac gtttacaccg acagccgcta tgcgttcgcg actgcccaca
     2161 tccacggtga aatctaccgc cgccgcggtt ggctgacgtc gaagggcaaa gaaatcaaaa
     2221 acaaagatga aatcttagct ctgctgaaag ctctgtttct gccgaaacgt ctgtccatta
     2281 ttcactgccc gggtcaccag aaagggcact ctgcagaagc tcgtggaaac cgtatggcag
     2341 accaggctgc gcgtaaagca gccatcactg aaaccccgga tacgtcaact ctgctgattg
     2401 aaaacagcag cccgaacagc cgcttgatta actgactcga ggtgatccgg ctgctaacaa
     2461 agcccgaaag gaagctgagt tggctgctgc caccgctgag caataactag cataacccct
     2521 tggggcctct aaacgggtct tgaggggttt tttgctgaaa ggaggaacta tatccggata
     2581 tcccgcaaga ggcccggcag taccggcata accaagccta tgcctacagc atccagggtg
     2641 acggtgccga ggatgacgat gagcgcattg ttagatttca tacacggtgc ctgactgcgt
     2701 tagcaattta actgtgataa actaccgcat taaagcttat cgatgataag ctgtcaaaca
     2761 tgagaattct tgaagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat
     2821 gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     2881 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     2941 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     3001 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     3061 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     3121 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     3181 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtgtt gacgccgggc aagagcaact
     3241 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     3301 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     3361 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     3421 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     3481 agccatacca aacgacgagc gtgacaccac gatgcctgca gcaatggcaa caacgttgcg
     3541 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     3601 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     3661 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
     3721 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     3781 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     3841 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     3901 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     3961 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     4021 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     4081 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     4141 accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     4201 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     4261 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     4321 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     4381 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     4441 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     4501 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     4561 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     4621 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     4681 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
     4741 cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt attttctcct
     4801 tacgcatctg tgcggtattt cacaccgcaa tggtgcactc tcagtacaat ctgctctgat
     4861 gccgcatagt taagccagta tacactccgc tatcgctacg tgactgggtc atggctgcgc
     4921 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg
     4981 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat
     5041 caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtga agcgattcac
     5101 agatgtctgc ctgttcatcc gcgtccagct cgttgagttt ctccagaagc gttaatgtct
     5161 ggcttctgat aaagcgggcc atgttaaggg cggttttttc ctgtttggtc actgatgcct
     5221 ccgtgtaagg gggatttctg ttcatggggg taatgatacc gatgaaacga gagaggatgc
     5281 tcacgatacg ggttactgat gatgaacatg cccggttact ggaacgttgt gagggtaaac
     5341 aactggcggt atggatgcgg cgggaccaga gaaaaatcac tcagggtcaa tgccagcgct
     5401 tcgttaatac agatgtaggt gttccacagg gtagccagca gcatcctgcg atgcagatcc
     5461 ggaacataat ggtgcagggc gctgacttcc gcgtttccag actttacgaa acacggaaac
     5521 cgaagaccat tcatgttgtt gctcaggtcg cagacgtttt gcagcagcag tcgcttcacg
     5581 ttcgctcgcg tatcggtgat tcattctgct aaccagtaag gcaaccccgc cagcctagcc
     5641 gggtcctcaa cgacaggagc acgatcatgc gcacccgtgg ccaggaccca acgctgcccg
     5701 agatgcgccg cgtgcggctg ctggagatgg cggacgcgat ggatatgttc tgccaagggt
     5761 tggtttgcgc attcacagtt ctccgcaaga attgattggc tccaattctt ggagtggtga
     5821 atccgttagc gaggtgccgc cggcttccat tcaggtcgag gtggcccggc tccatgcacc
     5881 gcgacgcaac gcggggaggc agacaaggta tagggcggcg cctacaatcc atgccaaccc
     5941 gttccatgtg ctcgccgagg cggcataaat cgccgtgacg atcagcggtc caatgatcga
     6001 agttaggctg gtaagagccg cgagcgatcc ttgaagctgt ccctgatggt cgtcatctac
     6061 ctgcctggac agcatggcct gcaacgcggg catcccgatg ccgccggaag cgagaagaat
     6121 cataatgggg aaggccatcc agcctcgcgt cgcgaacgcc agcaagacgt agcccagcgc
     6181 gtcggccgcc atgccggcga taatggcctg cttctcgccg aaacgtttgg tggcgggacc
     6241 agtgacgaag gcttgagcga gggcgtgcaa gattccgaat accgcaagcg acaggccgat
     6301 catcgtcgcg ctccagcgaa agcggtcctc gccgaaaatg acccagagcg ctgccggcac
     6361 ctgtcctacg agttgcatga taaagaagac agtcataagt gcggcgacga tagtcatgcc
     6421 ccgcgcccac cggaaggagc tgactgggtt gaaggctctc aagggcatcg gtcgagatcc
     6481 cggtgcctaa tgagtgagct aacttacatt aattgcgttg cgctcactgc ccgctttcca
     6541 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     6601 tttgcgtatt gggcgccagg gtggtttttc ttttcaccag tgagacgggc aacagctgat
     6661 tgcccttcac cgcctggccc tgagagagtt gcagcaagcg gtccacgctg gtttgcccca
     6721 gcaggcgaaa atcctgtttg atggtggtta acggcgggat ataacatgag ctgtcttcgg
     6781 tatcgtcgta tcccactacc gagatatccg caccaacgcg cagcccggac tcggtaatgg
     6841 cgcgcattgc gcccagcgcc atctgatcgt tggcaaccag catcgcagtg ggaacgatgc
     6901 cctcattcag catttgcatg gtttgttgaa aaccggacat ggcactccag tcgccttccc
     6961 gttccgctat cggctgaatt tgattgcgag tgagatattt atgccagcca gccagacgca
     7021 gacgcgccga gacagaactt aatgggcccg ctaacagcgc gatttgctgg tgacccaatg
     7081 cgaccagatg ctccacgccc agtcgcgtac cgtcttcatg ggagaaaata atactgttga
     7141 tgggtgtctg gtcagagaca tcaagaaata acgccggaac attagtgcag gcagcttcca
     7201 cagcaatggc atcctggtca tccagcggat agttaatgat cagcccactg acgcgttgcg
     7261 cgagaagatt gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct accatcgaca
     7321 ccaccacgct ggcacccagt tgatcggcgc gagatttaat cgccgcgaca atttgcgacg
     7381 gcgcgtgcag ggccagactg gaggtggcaa cgccaatcag caacgactgt ttgcccgcca
     7441 gttgttgtgc cacgcggttg ggaatgtaat tcagctccgc catcgccgct tccacttttt
     7501 cccgcgtttt cgcagaaacg tggctggcct ggttcaccac gcgggaaacg gtctgataag
     7561 agacaccggc atactctgcg acatcgtata acgttactgg tttcacattc accaccctga
     7621 attgactctc ttccgggcgc tatcatgcca taccgcgaaa ggttttgcgc cattcgatgg
     7681 tgtccgggat ctcgacgctc tcccttatgc gactcctgca ttaggaagca gcccagtagt
     7741 aggttgaggc cgttgagcac cgccgccgca aggaatggtg catgcaagg
//
