Subcellular localisation of marenostrin/pyrin isoforms carrying the most common mutations involved in familial Mediterranean fever in the presence or absence of its binding partner ASC -: art. no. e24
Subcellular localisation of marenostrin/pyrin isoforms carrying the most common mutations involved in familial Mediterranean fever in the presence or absence of its binding partner ASC -: art. no. e24
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DOI：
10.1136/jmg.2003.011601
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发表时间：
2004-03-01
影响因子：
4
通讯作者：
Amselem, S
中科院分区：
文献类型：
--
作者：
Cazeneuve, C;Papin, S;Amselem, S
METHODS Plasmid constructs The cDNAs coding for MEFV-fl and MEFV-d2 were obtained by digestion of the previously described pMEFV-fl-GFP and pMEFV-d2-GFP expression vectors11 by EcoRI and SalI. The resulting products corresponding to MEFV-fl and MEFV-d2 were then cloned into the pEGFP-N3 expression vector (Clontech), which had been previously modified by mutagenesis in order to inverse the SalI and EcoRI restriction sites of the polylinker. The resulting plasmids were named pEGFP-MEFV-fl and pEGFP-MEFV-d2. The M694V, M694I, M680I, V726A, E148Q, and A89T mutations were then introduced into pEGFP-MEFV-fl and pEGFP-MEFV-d2 using the Quick-Change site directed mutagenesis system (Stratagene). The cDNA encoding human ASC was generated from total leukocyte RNA, and amplified with forward primer 5!-CCCGCGTCGACGCAGCGGCCGGGGATCCTGGAGCC-3! and reverse primer 5!-GCCCAGAATTCCTCAGCTCCGCTCCAGGTCCTC-3!. The resulting RT-PCR product was subsequently cloned into pcDNA3. 1/V5-His-TOPO (Invitrogen), in frame with the V5 epitope, to generate the pASC-V5 expression plasmid.
