Flexitube control sirna

Mitochondrion-targeted DsRed2 (mtRFP) was previously described (Lee et al., 2012 ). Validated siRNAs (mtHsp40; Mm_Dnaja3_3 FlexiTube siRNA, mtHsp70; Hs_HSPA9_5 FlexiTube siRNA, Drp1; Hs_DNM1L_5 FlexiTube siRNA, and Scramble; FlexiTube Control siRNA) were purchased from Qiagen (Toronto, Canada). HA-Drp1 and Mff-Flag were kind gifts from Hidenori Otera (Kyushu University, Fukuoka, Japan). mtHsp70-Flag mutants were prepared from wild type purchased from OriGene (Rockville, MD). To generate mtHsp70-Flag mutants, PCR was performed using the following primer sequences: full length 1–679 (forward [FW]: 5′-BamHI ATG ATA AGT GCC AGC CGA GCT-3′; reverse [RV]: 5′-HindIII TTA GAG GTC TTC TTC TGA GAT GAG CTT CTG CTC CTG TTT TTC CTC CTT TTG AT-C TTC C-3′), ΔMTS 57-679 (FW: 5′-BamHI ATG ATA AGT GCC AGC CGA GCT-3′; RV: 5′-HindIII TTA GAG GTC TTC TTC TGA GAT GAG CTT CTG CTC CTG TTT TTC CTC CTT TTG ATC TTC C-3′), ΔATPase (FW1: 5′-BamHI ATG ATA AGT GCC AGC CGA GCT-3′; RV1: 5′- XbaI CAT AAC TGC CAC GCA GGA-3′; FW2: 5′-XbaI AGC TGC CAT TCA GGG AGG T-3′; RV2: 5′-HindIII TTA GAG GTC TTC TTC TGA GAT GAG CTT CTG CTC CTG TTT TTC CTC CTT TTG ATC TTC C-3′), and ΔCT 1–434 (FW: 5′-BamHI ATG ATA AGT GCC AGC CGA GCT-3′; RV: HindIII TTA GAG GTC TTC TTC TGA GAT GAG CTT CTG CTC ATC CGT GAC ATC GCC GGC CAA-3′).