pjnc-pgluc Gene/Insert name: Codon-optimized Gaussia luciferase (pgluc) Vector backbone: pcmv-jnc Vector type: Mammalian cells Backbone size w/o insert (bp): 5375 Bacterial resistance: Ampicillin and neomycin Growth strain: JM83 Growth temperature ( o C): 37 Growth instructions: pjnc-pgluc is resistant to ampicillin and neomycin High or low copy: High copy Vector map: pjnc-pgluc Coding sequence: Nucleotide sequence & Amino acid sequence Plasmid sequence: pjnc-pgluc (5,941 bp) Restriction enzyme list: Restriction enzyme sites of pjnc-pgluc GenBank Accession No.: - Size: 10 µg Constituents: 10 mm Tris-HCl (ph 7.6)-1 mm EDTA Terms and Licenses: MTA Laboratory Reagent For Research Use Only 1/6
Mammalian expression vector: Codon-optimized Gaussia Luciferase Name: pjnc-pgluc Insert: Codon-optimized Gaussia luciferase cdnas Vector: pcmv-jnc Amp r CMV promoter GLsp HindIII KpnI BamHI pgluc cdna puc origin pjnc-pgluc (5,941 bp) XbaI BGH poly A f1 origin SV40 promoter/ origin Feature for pjnc-pgluc: Residue 1-681 1-588 632-651 689-739 740-1,243 1,247-5,941 1,264-1,281 1,307-1,531 1,577-2,005 2,010-2,353 2,415-3,209 3,383-3,513 3,896-4,569 4,714-5,574 SV40 poly A Sourse 1-681 1-588 632-651 1-51 52-504 752-5,445 768-785 811-1,035 1,081-1,509 1,514-1,857 1,919-2,713 2,887-3,017 3,400-4,073 4,218-5,078 Neo r Comments pcmv-jnc backbone CMV promoter T7 promoter Signal peptide sequence of Gaussia luciferase pgluc ORF (K1-D168) pcmv-jnc backbone Sp6 promoter BGH polyadenylation sequence f1 origin SV40 early promoter and origin Neomycin resistance gene (ORF) SV40 early polyadenylation signal puc origin Ampicillin resistance gene (ORF) Ref. 1) pgluc: amino acid seq. & DNA seq.: GenBank Accession No. LC006266 2) Inouye, S. et al. Protein. Expr. Purif. (2015) 109: 47-54. 3) Inouye, S. et al. Protein. Expr. Purif. (2016) 128: 93-100. JNC Corporation, Yokohama Research Center
Gene coding region (ORF: Codon-optimized Gaussia luciferase) Nucleotide sequence GGATCCAGCCACCATGGGCGTCAAGGTCCTGTTCGCCCTGATCTGCATCGCCGTCGCCGAGGCCAAGCCC ACCGAGAACAACGAGGACTTCAACATCGTCGCCGTCGCCAGCAACTTCGCCACCACCGACCTGGACGCTG ACAGAGGCAAGCTGCCCGGCAAGAAGCTGCCCCTGGAGGTCCTGAAGGAGATGGAGGCCAACGCCAGAAA GGCCGGCTGCACCAGAGGCTGCCTGATCTGCCTGAGCCACATCAAGTGCACCCCCAAGATGAAGAAGTTC ATCCCCGGTAGATGCCACACCTACGAGGGCGACAAGGAGAGCGCCCAGGGCGGCATCGGCGAGGCCATCG TCGACATCCCCGAGATCCCCGGCTTCAAGGACCTGGAGCCCATGGAGCAGTTCATCGCCCAGGTCGACCT GTGCGTCGACTGCACCACCGGCTGCCTGAAGGGCCTGGCCAACGTCCAGTGCAGTGACCTGCTGAAGAAG TGGCTGCCCCAGAGATGCGCCACCTTCGCCAGCAAGATCCAGGGCCAGGTCGACAAGATCAAGGGCGCCG GCGGCGACTAATTCTAGA Amino acid sequence MGVKVLFALICIAVAEAKPTENNEDFNIVAVASNFATTDLDADRGKLPGKKLPLEVLKEMEANARKAGCT RGCLICLSHIKCTPKMKKFIPGRCHTYEGDKESAQGGIGEAIVDIPEIPGFKDLEPMEQFIAQVDLCVDC TTGCLKGLANVQCSDLLKKWLPQRCATFASKIQGQVDKIKGAGGD* 3/6
4/6 pjnc-pgluc (5,941 bp) GTTGACATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTCATAGCCCATATAT GGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCGCCCAACGACCCCCGCCCATTG ACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGGGACTTTCCATTGACGTCAATGGGTGGAGT ATTTACGGTAAACTGCCCACTTGGCAGTACATCAAGTGTATCATATGCCAAGTACGCCCCCTATTGACGT CAATGACGGTAAATGGCCCGCCTGGCATTATGCCCAGTACATGACCTTATGGGACTTTCCTACTTGGCAG TACATCTACGTATTAGTCATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACATCAATGGGCGTGGAT AGCGGTTTGACTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCA AAATCAACGGGACTTTCCAAAATGTCGTAACAACTCCGCCCCATTGACGCAAATGGGCGGTAGGCGTGTA CGGTGGGAGGTCTATATAAGCAGAGCTCTCTGGCTAACTAGAGAACCCACTGCTTACTGGCTTATCGAAA TTAATACGACTCACTATAGGGAGACCCAAGCTTGGTACCGAGCTCGGATCCAGCCACCATGGGCGTCAAG GTCCTGTTCGCCCTGATCTGCATCGCCGTCGCCGAGGCCAAGCCCACCGAGAACAACGAGGACTTCAACA TCGTCGCCGTCGCCAGCAACTTCGCCACCACCGACCTGGACGCTGACAGAGGCAAGCTGCCCGGCAAGAA GCTGCCCCTGGAGGTCCTGAAGGAGATGGAGGCCAACGCCAGAAAGGCCGGCTGCACCAGAGGCTGCCTG ATCTGCCTGAGCCACATCAAGTGCACCCCCAAGATGAAGAAGTTCATCCCCGGTAGATGCCACACCTACG AGGGCGACAAGGAGAGCGCCCAGGGCGGCATCGGCGAGGCCATCGTCGACATCCCCGAGATCCCCGGCTT CAAGGACCTGGAGCCCATGGAGCAGTTCATCGCCCAGGTCGACCTGTGCGTCGACTGCACCACCGGCTGC CTGAAGGGCCTGGCCAACGTCCAGTGCAGTGACCTGCTGAAGAAGTGGCTGCCCCAGAGATGCGCCACCT TCGCCAGCAAGATCCAGGGCCAGGTCGACAAGATCAAGGGCGCCGGCGGCGACTAATTCTAGAGGGCCCT ATTCTATAGTGTCACCTAAATGCTAGAGCTCGCTGATCAGCCTCGACTGTGCCTTCTAGTTGCCAGCCAT CTGTTGTTTGCCCCTCCCCCGTGCCTTCCTTGACCCTGGAAGGTGCCACTCCCACTGTCCTTTCCTAATA AAATGAGGAAATTGCATCGCATTGTCTGAGTAGGTGTCATTCTATTCTGGGGGGTGGGGTGGGGCAGGAC AGCAAGGGGGAGGATTGGGAAGACAATAGCAGGCATGCTGGGGATGCGGTGGGCTCTATGGCTTCTGAGG CGGAAAGAACCAGCTGGGGCTCTAGGGGGTATCCCCACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGG TGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTCTTC CCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCCCTTTAGGGTTCC GATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGGTGATGGTTCACGTAGTGGGCCATC GCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAA ACTGGAACAACACTCAACCCTATCTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCT ATTGGTTAAAAAATGAGCTGATTTAACAAAAATTTAACGCGAATTAATTCTGTGGAATGTGTGTCAGTTA GGGTGTGGAAAGTCCCCAGGCTCCCCAGCAGGCAGAAGTATGCAAAGCATGCATCTCAATTAGTCAGCAA CCAGGTGTGGAAAGTCCCCAGGCTCCCCAGCAGGCAGAAGTATGCAAAGCATGCATCTCAATTAGTCAGC AACCATAGTCCCGCCCCTAACTCCGCCCATCCCGCCCCTAACTCCGCCCAGTTCCGCCCATTCTCCGCCC CATGGCTGACTAATTTTTTTTATTTATGCAGAGGCCGAGGCCGCCTCTGCCTCTGAGCTATTCCAGAAGT AGTGAGGAGGCTTTTTTGGAGGCCTAGGCTTTTGCAAAAAGCTCCCGGGAGCTTGTATATCCATTTTCGG ATCTGATCAAGAGACAGGATGAGGATCGTTTCGCATGATTGAACAAGATGGATTGCACGCAGGTTCTCCG GCCGCTTGGGTGGAGAGGCTATTCGGCTATGACTGGGCACAACAGACAATCGGCTGCTCTGATGCCGCCG TGTTCCGGCTGTCAGCGCAGGGGCGCCCGGTTCTTTTTGTCAAGACCGACCTGTCCGGTGCCCTGAATGA ACTGCAGGACGAGGCAGCGCGGCTATCGTGGCTGGCCACGACGGGCGTTCCTTGCGCAGCTGTGCTCGAC GTTGTCACTGAAGCGGGAAGGGACTGGCTGCTATTGGGCGAAGTGCCGGGGCAGGATCTCCTGTCATCTC ACCTTGCTCCTGCCGAGAAAGTATCCATCATGGCTGATGCAATGCGGCGGCTGCATACGCTTGATCCGGC TACCTGCCCATTCGACCACCAAGCGAAACATCGCATCGAGCGAGCACGTACTCGGATGGAAGCCGGTCTT GTCGATCAGGATGATCTGGACGAAGAGCATCAGGGGCTCGCGCCAGCCGAACTGTTCGCCAGGCTCAAGG CGCGCATGCCCGACGGCGAGGATCTCGTCGTGACCCATGGCGATGCCTGCTTGCCGAATATCATGGTGGA AAATGGCCGCTTTTCTGGATTCATCGACTGTGGCCGGCTGGGTGTGGCGGACCGCTATCAGGACATAGCG TTGGCTACCCGTGATATTGCTGAAGAGCTTGGCGGCGAATGGGCTGACCGCTTCCTCGTGCTTTACGGTA TCGCCGCTCCCGATTCGCAGCGCATCGCCTTCTATCGCCTTCTTGACGAGTTCTTCTGAGCGGGACTCTG GGGTTCGAAATGACCGACCAAGCGACGCCCAACCTGCCATCACGAGATTTCGATTCCACCGCCGCCTTCT ATGAAAGGTTGGGCTTCGGAATCGTTTTCCGGGACGCCGGCTGGATGATCCTCCAGCGCGGGGATCTCAT GCTGGAGTTCTTCGCCCACCCCAACTTGTTTATTGCAGCTTATAATGGTTACAAATAAAGCAATAGCATC ACAAATTTCACAAATAAAGCATTTTTTTCACTGCATTCTAGTTGTGGTTTGTCCAAACTCATCAATGTAT CTTATCATGTCTGTATACCGTCGACCTCTAGCTAGAGCTTGGCGTAATCATGGTCATAGCTGTTTCCTGT GTGAAATTGTTATCCGCTCACAATTCCACACAACATACGAGCCGGAAGCATAAAGTGTAAAGCCTGGGGT GCCTAATGAGTGAGCTAACTCACATTAATTGCGTTGCGCTCACTGCCCGCTTTCCAGTCGGGAAACCTGT CGTGCCAGCTGCATTAATGAATCGGCCAACGCGCGGGGAGAGGCGGTTTGCGTATTGGGCGCTCTTCCGC
TTCCTCGCTCACTGACTCGCTGCGCTCGGTCGTTCGGCTGCGGCGAGCGGTATCAGCTCACTCAAAGGCG GTAATACGGTTATCCACAGAATCAGGGGATAACGCAGGAAAGAACATGTGAGCAAAAGGCCAGCAAAAGG CCAGGAACCGTAAAAAGGCCGCGTTGCTGGCGTTTTTCCATAGGCTCCGCCCCCCTGACGAGCATCACAA AAATCGACGCTCAAGTCAGAGGTGGCGAAACCCGACAGGACTATAAAGATACCAGGCGTTTCCCCCTGGA AGCTCCCTCGTGCGCTCTCCTGTTCCGACCCTGCCGCTTACCGGATACCTGTCCGCCTTTCTCCCTTCGG GAAGCGTGGCGCTTTCTCATAGCTCACGCTGTAGGTATCTCAGTTCGGTGTAGGTCGTTCGCTCCAAGCT GGGCTGTGTGCACGAACCCCCCGTTCAGCCCGACCGCTGCGCCTTATCCGGTAACTATCGTCTTGAGTCC AACCCGGTAAGACACGACTTATCGCCACTGGCAGCAGCCACTGGTAACAGGATTAGCAGAGCGAGGTATG TAGGCGGTGCTACAGAGTTCTTGAAGTGGTGGCCTAACTACGGCTACACTAGAAGAACAGTATTTGGTAT CTGCGCTCTGCTGAAGCCAGTTACCTTCGGAAAAAGAGTTGGTAGCTCTTGATCCGGCAAACAAACCACC GCTGGTAGCGGTGGTTTTTTTGTTTGCAAGCAGCAGATTACGCGCAGAAAAAAAGGATCTCAAGAAGATC CTTTGATCTTTTCTACGGGGTCTGACGCTCAGTGGAACGAAAACTCACGTTAAGGGATTTTGGTCATGAG ATTATCAAAAAGGATCTTCACCTAGATCCTTTTAAATTAAAAATGAAGTTTTAAATCAATCTAAAGTATA TATGAGTAAACTTGGTCTGACAGTTACCAATGCTTAATCAGTGAGGCACCTATCTCAGCGATCTGTCTAT TTCGTTCATCCATAGTTGCCTGACTCCCCGTCGTGTAGATAACTACGATACGGGAGGGCTTACCATCTGG CCCCAGTGCTGCAATGATACCGCGAGACCCACGCTCACCGGCTCCAGATTTATCAGCAATAAACCAGCCA GCCGGAAGGGCCGAGCGCAGAAGTGGTCCTGCAACTTTATCCGCCTCCATCCAGTCTATTAATTGTTGCC GGGAAGCTAGAGTAAGTAGTTCGCCAGTTAATAGTTTGCGCAACGTTGTTGCCATTGCTACAGGCATCGT GGTGTCACGCTCGTCGTTTGGTATGGCTTCATTCAGCTCCGGTTCCCAACGATCAAGGCGAGTTACATGA TCCCCCATGTTGTGCAAAAAAGCGGTTAGCTCCTTCGGTCCTCCGATCGTTGTCAGAAGTAAGTTGGCCG CAGTGTTATCACTCATGGTTATGGCAGCACTGCATAATTCTCTTACTGTCATGCCATCCGTAAGATGCTT TTCTGTGACTGGTGAGTACTCAACCAAGTCATTCTGAGAATAGTGTATGCGGCGACCGAGTTGCTCTTGC CCGGCGTCAATACGGGATAATACCGCGCCACATAGCAGAACTTTAAAAGTGCTCATCATTGGAAAACGTT CTTCGGGGCGAAAACTCTCAAGGATCTTACCGCTGTTGAGATCCAGTTCGATGTAACCCACTCGTGCACC CAACTGATCTTCAGCATCTTTTACTTTCACCAGCGTTTCTGGGTGAGCAAAAACAGGAAGGCAAAATGCC GCAAAAAAGGGAATAAGGGCGACACGGAAATGTTGAATACTCATACTCTTCCTTTTTCAATATTATTGAA GCATTTATCAGGGTTATTGTCTCATGAGCGGATACATATTTGAATGTATTTAGAAAAATAAACAAATAGG GGTTCCGCGCACATTTCCCCGAAAAGTGCCACCTGACGTCGACGGATCGGGAGATCTCCCGATCCCCTAT GGTGCACTCTCAGTACAATCTGCTCTGATGCCGCATAGTTAAGCCAGTATCTGCTCCCTGCTTGTGTGTT GGAGGTCGCTGAGTAGTGCGCGAGCAAAATTTAAGCTACAACAAGGCAAGGCTTGACCGACAATTGCATG AAGAATCTGCTTAGGGTTAGGCGTTTTGCGCTGCTTCGCGATGTACGGGCCAGATATACGC Residue Source Comments 1-681 1-681 pcmv-jnc backbone (pcdna3 derivative) 1-588 1-588 CMV promoter 632-651 632-651 T7 promoter 689-739 1-51 Signal peptide sequence of Gaussia luciferase 740-1,243 52-504 pgluc ORF (K1-D168) 1,247-5,941 752-5,445 pcmv-jnc backbone (pcdna3 derivative) 1,264-1,281 768-785 Sp6 promoter 1,307-1,531 811-1,035 BGH polyadenylation sequence 1,577-2,005 1,081-1,509 f1 origin 2,010-2,353 1,514-1,857 SV40 early promoter and origin 2,415-3,209 1,919-2,713 Neomycin resistance gene (ORF) 3,383-3,513 2,887-3,017 SV40 early polyadenylation signal 3,896-4,569 3,400-4,073 puc origin 4,714-5,574 4,218-5,078 Ampicillin resistance gene (ORF) 5/6
Restriction enzyme sites of pjnc-pgluc Enzyme Name Sequence Count Cutting Positions AccI GT!MKAC 7 1027, 1090, 1102, 1216, 3515, 3522, 5710 ApaI GGGCC!C 1 1259 Asp718I G!GTACC 1 665 BamHI G!GATCC 1 677 BclI T!GATCA 2 1295, 2385 BglII A!GATCT 1 5723 EcoRI G!AATTC 0 - EcoRV GAT!ATC 0 - HincII GTY!RAC 7 4, 1028, 1091, 1103, 1217,3523, 5711 HindIII A!AGCTT 1 659 KpnI GGTAC!C 1 669 MluI A!CGCGT 0 - NcoI C!CATGG 5 380, 688, 1066, 2241, 2976 NdeI CA!TATG 1 254 NheI G!CTAGC 0 - NotI GC!GGCCGC 0 - PstI CTGCA!G 1 2597 SacI GAGCT!C 3 588, 675, 1291 SalI G!TCGAC 6 1026, 1089, 1101, 1215, 3521, 5709 ScaI AGT!ACT 1 5268 SmaI CCC!GGG 1 2357 StuI AGG!CCT 1 2333 XbaI T!CTAGA 1 1249 XhoI C!TCGAG 0 - Supplier Shin Otemachi Bldg. 9F 2-2-1 Otemachi, Chiyoda-ku, Tokyo 100-8105 URL http://www.jnc-corp.co.jp Contact us JNC Corporation, Yokohama Research Center 5-1 Okawa, Kanazawa-ku, Yokohama, Japan 236-8605 Tel: 045-786-5501 Fax: 045-786-5511 E-mail: biophoton@jnc-corp.co.jp 6/6