LOCUS       sequence-331338-        5150 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of Nsp9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5150)
  TITLE     Savchenko Lab Plasmids
REFERENCE   2  (bases 1 to 5150)
  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: pMCSG53_Nsp9
FEATURES             Location/Qualifiers
     source          1..5150
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     940..959
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(979..1056)
                     /gene="lacI"
                     /label=lacI promoter
     tRNA            1196..1271
                     /gene="ileX"
                     /product="E. coli tRNA-Ile recognizing AUA"
                     /label=ileX
     primer_bind     1420..1439
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     tRNA            1541..1617
                     /gene="argU"
                     /product="E. coli tRNA-Arg recognizing AGA and AGG"
                     /label=argU
     primer_bind     1892..1911
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1892..1910
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1911..1935
                     /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             1950..1972
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1983..2000
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2025..2045
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2456..2473
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2526..2544)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2540..2587
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(2711..2730)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2897..3000
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3001..3069,3070..3861)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3219..3238)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4032..4620
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4521..4540
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4774..4791
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(4871..4890)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             complement(join(5046..5150,1..978))
                     /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 tcttttcacc agtgagacgg gcaacagctg attgcccttc accgcctggc cctgagagag
       61 ttgcagcaag cggtccacgc tggtttgccc cagcaggcga aaatcctgtt tgatggtggt
      121 taacggcggg atataacatg agctgtcttc ggtatcgtcg tatcccacta ccgagatatc
      181 cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg ccatctgatc
      241 gttggcaacc agcatcgcag tgggaacgat gccctcattc agcatttgca tggtttgttg
      301 aaaaccggac atggcactcc agtcgccttc ccgttccgct atcggctgaa tttgattgcg
      361 agtgagatat ttatgccagc cagccagacg cagacgcgcc gagacagaac ttaatgggcc
      421 cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc ccagtcgcgt
      481 accgtcttca tgggagaaaa taatactgtt gatgggtgtc tggtcagaga catcaagaaa
      541 taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt catccagcgg
      601 atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg ccgctttaca
      661 ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca gttgatcggc
      721 gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac tggaggtggc
      781 aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt tgggaatgta
      841 attcagctcc gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa cgtggctggc
      901 ctggttcacc acgcgggaaa cggtctgata agagacaccg gcatactctg cgacatcgta
      961 taacgttact ggtttcacat tcaccaccct gaattgactc tcttccgggc gctatcatgc
     1021 cataccgcga aaggttttgc gccattcgat ggtgtccggg atctcgacgc tctcccttat
     1081 gcgactcctg cattaggaag cagcccagta gtaggttgag gccgttgagc accgccgccg
     1141 caaggaatgg tgcatgcgga ttgcgacacg gagttacttt ataatccgct accatggccc
     1201 cttagctcag tggttagagc aggcgactca taatcgcttg gtcgctggtt caagtccagc
     1261 aggggccacc agatatagca aaggctgacg agaaatcgtc agcctttttg catgcaagga
     1321 gatggcgccc aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc
     1381 gctcatgagc ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc
     1441 gccagcaacc gcacctgtgg cgccggcgag atcaaaagcc attgactcag caagggttga
     1501 ccgtataatt cacgcgatta caccgcattg cggtatcaac gcgcccttag ctcagttgga
     1561 tagagcaacg accttctaag tcgtgggccg caggttcgaa tcctgcaggg cgcgccatta
     1621 caattcaatc agttacgcct tctttatatc ctccataatt ccagagtggg acatatttgg
     1681 gacattatca ccaaaaatgt cgtctatttt cctcgcatgc tctgtcaaat gattaggcgc
     1741 aaggtgagca tacctacgaa ccatttctat ggactcccat ccgcccattt cctgaagcac
     1801 tgataatggg acgcctgact gaatccaccg gtgatgccgg ccacgatgcg tccggcgtag
     1861 aggatcgaga tcgatctcga tcccgcgaaa ttaatacgac tcactatagg ggaattgtga
     1921 gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga gatatacata
     1981 tgcaccatca tcatcatcat tcttctggtg tagatctggg taccgagaac ctgtacttcc
     2041 aatccaatat gaacaatgaa ttatcacccg tagctctaag gcaaatgagc tgcgctgccg
     2101 gtactacgca gaccgcatgc accgatgata atgcgctggc gtattacaat accaccaaag
     2161 gcggtcgttt cgtgttggcg ttgctgtccg acctccaaga tttaaaatgg gcacgttttc
     2221 cgaagtctga cggcacgggt acgatttaca ccgaactgga gccgccatgt cgcttcgtca
     2281 ccgacacccc gaagggtccg aaggtgaaat acctgtattt tatcaaaggc ctgaacaacc
     2341 tgaaccgcgg tatggttttg ggcagcctgg cggctaccgt tcgtctgcag taaattggaa
     2401 gtggataacg gatccgaatt cgagctccgt cgacaagctt gcggccgcac tcgagcacca
     2461 ccaccaccac cactgagatc cggctgctaa caaagcccga aaggaagctg agttggctgc
     2521 tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg tcttgagggg
     2581 ttttttgctg aaaggaggaa ctatatccgg attggcgaat gggacgcgcc ctgtagcggc
     2641 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     2701 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     2761 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     2821 gaccccaaaa aacttgatta gggtgatggt tcacattaac gcttacaatt taggtggcac
     2881 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     2941 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatttgaaa aaggaagagt
     3001 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     3061 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     3121 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     3181 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     3241 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     3301 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     3361 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     3421 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     3481 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     3541 cctgcagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     3601 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     3661 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     3721 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     3781 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     3841 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     3901 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     3961 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     4021 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     4081 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     4141 gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta
     4201 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     4261 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     4321 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     4381 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     4441 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     4501 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     4561 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     4621 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     4681 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     4741 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     4801 gagcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcaatggt
     4861 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc
     4921 gctacgtgac tgtagtcatg ccccgcgccc accggaagga gctgactggg ttgaaggctc
     4981 tcaagggcat cggtcgagat cccggtgcct aatgagtgag ctaacttaca ttaattgcgt
     5041 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     5101 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca gggtggtttt
//
