RU2018141183A - Recombinant plasmid DNA pET32a-NS1-ZIKV, which provides expression of the NS1 gene encoding the non-structural protein Z1 virus NS1 and recombinant protein NS1-ZIKV, designed to induce specific immunity against Zika virus - Google Patents

Recombinant plasmid DNA pET32a-NS1-ZIKV, which provides expression of the NS1 gene encoding the non-structural protein Z1 virus NS1 and recombinant protein NS1-ZIKV, designed to induce specific immunity against Zika virus Download PDF

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RU2018141183A
RU2018141183A RU2018141183A RU2018141183A RU2018141183A RU 2018141183 A RU2018141183 A RU 2018141183A RU 2018141183 A RU2018141183 A RU 2018141183A RU 2018141183 A RU2018141183 A RU 2018141183A RU 2018141183 A RU2018141183 A RU 2018141183A
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zikv
protein
recombinant
gene
expression
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RU2018141183A
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Russian (ru)
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RU2018141183A3 (en
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Елена Ивановна Казачинская
Наталья Вячеславовна Волкова
Даниил Васильевич Шаньшин
Анна Владимировна Зайковская
Алла Владимировна Иванова
Дмитрий Николаевич Щербаков
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Федеральное бюджетное учреждение науки "Государственный научный центр вирусологии и биотехнологии "Вектор" Федеральной службы по надзору в сфере защиты прав потребителей и благополучия человека (ФБУН ГНЦ ВБ "Вектор" Роспотребнадзора)
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Priority to RU2018141183A priority Critical patent/RU2018141183A/en
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Publication of RU2018141183A3 publication Critical patent/RU2018141183A3/ru

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Claims (11)

1. Рекомбинантная плазмидная ДНК pЕТ32a-NS1-ZIKV, обеспечивающая экспрессию гена NS1, кодирующего неструктурный белок NS1 вируса Зика, предназначенная для получения рекомбинантного белка NS1-ZIKV, обеспечивающего индукцию специфического иммунитета против вируса Зика, имеющая размер 6982 п.н. и содержащая в своем составе: 1. Recombinant plasmid DNA pET32a-NS1-ZIKV, providing the expression of the NS1 gene encoding a non-structural protein NS1 Zika virus, designed to obtain a recombinant protein NS1-ZIKV, providing the induction of specific immunity against Zika virus, having a size of 6982 bp and containing in its composition: - ген bla (координаты с 599 по 1459 п.н), в качестве генетического маркера, определяющего устойчивость к ампициллину клеток бактерии E.coli, трансформированных рекомбинантной плазмидой pЕТ32a- NS1-ZIKV;- bla gene (coordinates from 599 to 1459 bp), as a genetic marker that determines ampicillin resistance of E. coli bacteria cells transformed with the recombinant plasmid pET32a- NS1-ZIKV; - участок начала репликации ori (координаты с 1630 по 2218 п.н.)- Ori replication start site (coordinates from 1630 to 2218 bp) - промотор фага T7 (координаты с 5121 по 5139 п.н.), обеспечивающий экспрессию последовательности гена NS1 ZIKV в прокариотических клетках E.coli;- T7 phage promoter (coordinates 5121 to 5139 bp), which provides expression of the ZIKV NS1 gene sequence in prokaryotic E. coli cells; - лактозный репрессор (координаты с 3648 п.н. по 4730 п.н.) - ДНК - связывающий белок, который ингибирует экспрессию генов E.coli и обеспечивает экспрессию целевого рекомбинантного белка после добавления индуктора изопропил-β-D-1-тиогалактопиранозид (IPTG);- lactose repressor (coordinates from 3648 bp to 4730 bp) - DNA - a binding protein that inhibits the expression of E. coli genes and ensures the expression of the target recombinant protein after the addition of the isopropyl-β-D-1-thiogalactopyranoside inducer ( IPTG); - уникальные сайты узнавания рестрикционными эндонуклеазами, имеющие следующие координаты: SalI (6799 п.н.) и EcoRI (5704 п.н.);- Unique recognition sites by restriction endonucleases having the following coordinates: SalI (6799 bp) and EcoRI (5704 bp); - ген NS1 ZIKV (1113 п.н.), встроенный в экспрессионную кассету по сайтам рестрикции SalI (6799 п.н.) и EcoRI (5704 п.н.), который кодирует 371 аминокислотных остатков (а.о.) нативного секреторного гликопротеина NS1;- NS1 ZIKV gene (1113 bp), built into the expression cassette at the restriction sites SalI (6799 bp) and EcoRI (5704 bp), which encodes 371 amino acid residues (aa) of the native secretory glycoprotein NS1; - ген тиоредоксина E.coli (с 5209 п.н. по 5535 п.н.) (327 п.н.), расположенный перед фрагментом гена NS1, кодирующего неструктурный белок NS1 ZIKV;- E.coli thioredoxin gene (from 5209 bp to 5535 bp) (327 bp), located in front of the fragment of the NS1 gene encoding the non-structural protein NS1 ZIKV; - последовательность (18 п.н.), кодирующая полигистидиновый (6xHis) политракт из шести гистидинов на N-конце кодируемого белка для последующей очистки его с помощью метода аффинной никель - хроматографии. - a sequence (18 bp) encoding a polyhistidine (6xHis) poly tract of six histidines at the N-terminus of the encoded protein for subsequent purification using the method of affinity nickel chromatography. 2. Рекомбинантный белок NS1-ZIKV, полученный в результате экспрессии гена NS1 вируса Зика в бактериальной системе E.coli в составе рекомбинантной плазмидной ДНК pЕТ32a-NS1-ZIKV по п. 1 формулы, имеющий молекулярную массу 60,9 кДа и аминокислотную последовательность SEQ ID NO:1 длиной 1088 а.к.о.:2. Recombinant NS1-ZIKV protein obtained by expression of the Zika virus NS1 gene in the E. coli bacterial system as part of the recombinant pET32a-NS1-ZIKV plasmid DNA according to claim 1, having a molecular weight of 60.9 kDa and the amino acid sequence of SEQ ID NO: 1, length 1088 a.c.o .: GTGGGGTGCTCGGTGGACTTCTCAAAGAAGGAGACGAGATGCGGTACAGGGGTGTTCGTCTATAACGACGTTGAAGCCTGGAGGGACAGGTACAAGTACCATCCTGACTCCCCCCGTAGATTGGCAGCAGCAGTCAAGCAAGCCTGGGAAGATGGTATCTGCGGGATCTCCTCTGTTTCAAGAATGGAAAACATCATGTGGAGATCAGTAGAAGGGGAGCTCAACGCAATCCTGGAAGAGAATGGAGTTCAACTGACGGTCGTTGTGGGATCTGTAAAAAACCCCATGTGGGGAGGTCCACAGAGATTGCCCGTGCCTGTGAACGAGCTGCCCCACGGCTGGAAGGCTTGGGGGAAATCGCACTTCGTCAGAGCAGCAAAGACAAATAACAGCTTTGTCGTGGATGGTGACACACTGAAGGAATGCCCACTCAAACATAGAGCATGGAACAGCTTTCTTGTGGAGGATCATGGGTTCGGGGTATTTCACACTAGTGTCTGGCTCAAGGTTAGAGAAGATTATTCATTAGAGTGTGATCCAGCCGTTATTGGAACAGCTGTTAAGGGAAAGGAGGCTGTACACAGTGATCTAGGCTACTGGATTGAGAGTGAGAAGAATGACACATGGAGGCTGAAGAGGGCCCATCTGATCGAGATGAAAACATGTGAATGGCCAAAGTCCCACACATTGTGGACAGATGGAATAGAAGAGAGTGATCTGATCATACCCAAGTCTTTAGCTGGGCCACTCAGCCATCACAATACCAGAGAGGGCTACAGGACCCAAATGAAAGGGCCATGGCACAGTGAAGAGCTTGAAATTCGGTTTGAGGAATGCCCAGGCACTAAGGTCCACGTGGAGGAAACATGTGGAACAAGAGGACCATCTCTGAGATCAACCACTGCAAGCGGAAGGGTGATCGAGGAATGGTGCTGCAGGGAGTGCACAATGCCCCCACTGTCGTTCCGGGCTAAAGATGGCTGTTGGTATGGAATGGAGATAAGGCCCAGGAAAGAACCAGAAAGCAACTTAGTAAGGTCAATGGTGACTGCAGGATCAACTGATCACATGGATCACTTCTCCCTTGG, обладающий антигенными и иммуногенными свойствами и предназначенный для индукции специфического иммунитета против вируса Зика.GTGGGGTGCTCGGTGGACTTCTCAAAGAAGGAGACGAGATGCGGTACAGGGGTGTTCGTCTATAACGACGTTGAAGCCTGGAGGGACAGGTACAAGTACCATCCTGACTCCCCCCGTAGATTGGCAGCAGCAGTCAAGCAAGCCTGGGAAGATGGTATCTGCGGGATCTCCTCTGTTTCAAGAATGGAAAACATCATGTGGAGATCAGTAGAAGGGGAGCTCAACGCAATCCTGGAAGAGAATGGAGTTCAACTGACGGTCGTTGTGGGATCTGTAAAAAACCCCATGTGGGGAGGTCCACAGAGATTGCCCGTGCCTGTGAACGAGCTGCCCCACGGCTGGAAGGCTTGGGGGAAATCGCACTTCGTCAGAGCAGCAAAGACAAATAACAGCTTTGTCGTGGATGGTGACACACTGAAGGAATGCCCACTCAAACATAGAGCATGGAACAGCTTTCTTGTGGAGGATCATGGGTTCGGGGTATTTCACACTAGTGTCTGGCTCAAGGTTAGAGAAGATTATTCATTAGAGTGTGATCCAGCCGTTATTGGAACAGCTGTTAAGGGAAAGGAGGCTGTACACAGTGATCTAGGCTACTGGATTGAGAGTGAGAAGAATGACACATGGAGGCTGAAGAGGGCCCATCTGATCGAGATGAAAACATGTGAATGGCCAAAGTCCCACACATTGTGGACAGATGGAATAGAAGAGAGTGATCTGATCATACCCAAGTCTTTAGCTGGGCCACTCAGCCATCACAATACCAGAGAGGGCTACAGGACCCAAATGAAAGGGCCATGGCACAGTGAAGAGCTTGAAATTCGGTTTGAGGAATGCCCAGGCACTAAGGTCCACGTGGAGGAAACATGTGGAACAAGAGGACCATCTCTGAGATCAACCACTGCAAGCGGAAGGGTGATCGAGGAATGGTGCTGCAGGGAGTGCACAATGCCCCCACTGTCGTTCCGGGCTAAAGATGGCTGTTGGTATGGAATGGAGA TAAGGCCCAGGAAAGAACCAGAAAGCAACTTAGTAAGGTCAATGGTGACTGCAGGATCAACTGATCACATGGATCACTTCTCCCTTGG, which has antigenic and immunogenic properties and is intended to induce specific immunity against Zika virus.
RU2018141183A 2018-11-22 2018-11-22 Recombinant plasmid DNA pET32a-NS1-ZIKV, which provides expression of the NS1 gene encoding the non-structural protein Z1 virus NS1 and recombinant protein NS1-ZIKV, designed to induce specific immunity against Zika virus RU2018141183A (en)

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