LOCUS Exported 4707 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp10. ACCESSION . VERSION . KEYWORDS pGBW-m4133845 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4707) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 4707) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4707 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 425..445 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 446..523 /codon_start=1 /product="calmodulin-binding peptide" /label=CBP /note="derived from skeletal muscle myosin light chain kinase; binds calmodulin with nanomolar affinity in the presence of calcium" /translation="KRRWKKNFIAVSAANRFKKISSSGAL" polyA_signal 576..697 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(587..606) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 641..660 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 812..858 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 888..1024 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 1069..1140 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(1366..1954) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1446..1465) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS 2700..3560 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind complement(2918..2937) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 3573..3709 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 3861..3932 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 3937..3983 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 4001..4049 /note="synthetic polyadenylation signal" enhancer 4130..4433 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 4435..4637 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 4588..4608 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 4706..8 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" ORIGIN 1 caccatggca gggaatgcga cagaggttcc tgccaactca acggtcttgt ctttttgtgc 61 tttcgccgtg gacgctgcaa aagcctacaa ggattatcta gcctcgggtg gacagccaat 121 cactaactgc gtaaagatgc tttgtaccca caccggcact ggccaagcaa ttacagtgac 181 ccccgaagct aatatggacc aggagtcctt cggaggtgcg agctgctgcc tgtactgtcg 241 ttgtcatata gatcacccga atcccaaagg gttttgcgac ctcaagggca aatatgtcca 301 gatcccaacg acatgcgcca acgatcctgt gggattcact ctgaagaaca ccgtatgtac 361 cgtttgcggg atgtggaagg ggtatggctg tagttgcgac caattacggg aaccgatgct 421 gcaagaaaac ctttactttc agtcgaagag gcgatggaaa aagaatttta tcgcggtgag 481 tgcagccaac agattcaaaa agatatcctc gtctggtgcc ctctaagcta gcttgactga 541 ctgagataca gcgtaccttc agctcacaga catgataaga tacattgatg agtttggaca 601 aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc 661 tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt 721 tatgtttcag gttcaggggg aggtgtggga ggttttttaa agcaagtaaa acctctacaa 781 atgtggtatt ggcccatctc tatcggtatc gtagcataac cccttggggc ctctaaacgg 841 gtcttgaggg gttttttgtg cccctcgggc cggattgcta tctaccggca ttggcgcaga 901 aaaaaatgcc tgatgcgacg ctgcgcgtct tatactccca catatgccag attcagcaac 961 ggatacggct tccccaactt gcccacttcc atacgtgtcc tccttaccag aaatttatcc 1021 ttaaggtcgt cagctatcct gcaggcgatc tctcgatttc gatcaagaca ttcctttaat 1081 ggtcttttct ggacaccact aggggtcaga agtagttcat caaactttct tccctcccta 1141 atctcattgg ttaccttggg ctatcgaaac ttaattaacc agtcaagtca gctacttggc 1201 gagatcgact tgtctgggtt tcgactacgc tcagaattgc gtcagtcaag ttcgatctgg 1261 tccttgctat tgcacccgtt ctccgattac gagtttcatt taaatcatgt gagcaaaagg 1321 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg 1381 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg 1441 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac 1501 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca 1561 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt 1621 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc 1681 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag 1741 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac 1801 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt 1861 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa 1921 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg 1981 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa 2041 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat 2101 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc 2161 gatctgtcta tttcgttcat ccatagttgc atttaaattt ccgaactctc caaggccctc 2221 gtcggaaaat cttcaaacct ttcgtccgat ccatcttgca ggctacctct cgaacgaact 2281 atcgcaagtc tcttggccgg ccttgcgcct tggctattgc ttggcagcgc ctatcgccag 2341 gtattactcc aatcccgaat atccgagatc gggatcaccc gagagaagtt caacctacat 2401 cctcaatccc gatctatccg agatccgagg aatatcgaaa tcggggcgcg cctggtgtac 2461 cgagaacgat cctctcagtg cgagtctcga cgatccatat cgttgcttgg cagtcagcca 2521 gtcggaatcc agcttgggac ccaggaagtc caatcgtcag atattgtact caagcctggt 2581 cacggcagcg taccgatctg tttaaaccta gatattgata gtctgatcgg tcaacgtata 2641 atcgagtcct agcttttgca aacatctatc aagagacagg atcagcagga ggctttcgca 2701 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg 2761 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcgc 2821 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg 2881 aagaacgctt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc 2941 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg 3001 ttgagtattc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat 3061 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgattg 3121 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg 3181 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc 3241 ctgtagcaat ggcaacaacc ttgcgtaaac tattaactgg cgaactactt actctagctt 3301 cccggcaaca gttgatagac tggatggagg cggataaagt tgcaggacca cttctgcgct 3361 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc 3421 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca 3481 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct 3541 cactgattaa gcattggtaa ccgattctag gtgcattggc gcagaaaaaa atgcctgatg 3601 cgacgctgcg cgtcttatac tcccacatat gccagattca gcaacggata cggcttcccc 3661 aacttgccca cttccatacg tgtcctcctt accagaaatt tatccttaag atcccgaatc 3721 gtttaaactc gactctggct ctatcgaatc tccgtcgttt cgagcttacg cgaacagccg 3781 tggcgctcat ttgctcgtcg ggcatcgaat ctcgtcagct atcgtcagct tacctttttg 3841 gcagcgatcg cggctcccga catcttggac cattagctcc acaggtatct tcttccctct 3901 agtggtcata acagcagctt cagctacctc tcaattcaaa aaacccctca agacccgttt 3961 agaggcccca aggggttatg ctatcaatcg ttgcgttaca cacacaaaaa accaacacac 4021 atccatcttc gatggatagc gattttatta tctaactgct gatcgagtgt agccagatct 4081 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac 4141 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa 4201 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt 4261 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc 4321 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat 4381 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atgctgatgc 4441 ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc 4501 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa 4561 aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg 4621 tctatataag cagagctggt ttagtgaacc gtcagatcag atctttgtcg atcctaccat 4681 ccactcgaca cacccgccag cggccgc //