Control:Article Title: RENEB Inter-Laboratory Comparison 2021: The Gene Expression Assay
Article Snippet: .. Additionally, all column RNA prep kits remove most of the DNA. (−) RT control conventional PCR (ß-actin primer, HotStar MasterMix (Qiagen), 30 cycles) Check DNA conta mination No cDNA synthesis cDNA synthesis Kit/MasterMix High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) QuantiTect Reverse Transcription (Qiagen) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) RevertAid First Strand cDNA Synthesis Kit (Thermo Scientific) High Capacity cDNA Archive Kit High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) Kit/MasterMix Quick Amp Labeling Kit (Agilent) PCR protocol 1× /25°C/10min, 1×/37°C/ 120min, 1×/85°C/5min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/42°C/2min, 1×/42°C/20min, 1×/95°C/3min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/25°C/5min, 1×/42°C/60min, 1×/l0°C/5min 1×/25°C/10min, 1×/37°C/120min, 1× /85°C/5min 1×/25°C/10min, 1×/37°C/120min PCR protocol 1×/40°C/120min, 1×/70°C/15min; 1 × /40°C/120min Quality control UBC Ct ITFG1 Ct, DPM1 Ct MRPS5 Ct No HPRT1 Ct 18S rRNA Ct Quality control NanoDrop TM qRT-PCR Kit/MasterMix TaqMan Universal Master Mix TaqMan Universal Master Mix II, no UNG (Thermo Fisher Scientific) QuantiFast SYBR Green PCR (Qiagen) 5X HOT FIREPol ® EvaGreen ® qPCR SuperMix, Solis BioDyne TaqMan fast advanced master mix (Applied Biosystems) and Maxima SYBR Green qPCR Master Mix (Thermo Scientific) TaqMan,PerfeCTa ® , MultiPlex qPCR SuperMix, Quanta bioscience TaqMan Universal Master Mix Microarray DNA-Microarray Agilent, 44k whole human genome, G4112F TaqMan assays SYBR Green assay FDXR (Hs00244586_ml), GDF15 (Hs00171132_ml) BAX (Hs00180269_ml), BBC3 (Hs00248075_ml), CDKN1A (Hs00355782_ml), DDB2 (Hs03044953_ml), FDXR (Hs00244586_ml), GADD45A (Hs00169255_ml), GDF15 (Hs00171132_ml), TNFSF4 (Hs00182411_ml) CDKN1A-F: AGACCAGCATGACAGATTTCTACC; CDKN1A-R: CTTCCTGTGGGCGGATTAGG; DDB2-F: AGCATCACTGGGCTGAAGTT; DDB2-R: TGGTGTCTGAGCTGGCAAAA; FDX-F: TGGAGAGAACGGACATCACG; FDX-R: AGCCACACTGTCTTCACTCG GADD45a for: ACTGCGTGCTGGTGACGAAT, GADD45a rev: GTTGACTTAAGGCAGGATCCTTCCA; FDXR for: TGGATGTGCCAGGCCTCTAC, FDXR rev: TGAGGAAGCTGTCAGTCATGGTT; CDKN1A for: CCTGGAGACTCTCAGGGTCGAAA, CDKN1A rev: GCGTTTGGAGTGGTAGAAATCTGTCA; MDM2 for: TATCAGGCAGGGGAGAGTGATACA, MDM2 rev: CCAACATCTGTTGCAATGTGATGGAA; 18S for: GCTTAATTTGACTCAACACGGGA, 18S rev: AGCTATCAATCTGTCAATCCTGTCC. .. TaqMan ® assay: DDB2 (Hs00172068_ml), FDXR (HS01031617_ml), ITFG1: Hs01061271_ml SYBR Green assay: CDKN1A F:CCT CAT CCC GTG TTC TCC TTT CDKN1A R: GTA CCA CCC AGC GGA CAA GT GAPDH F: CGA CCA CTT TGT CAA GCT CA GAPDH R: AGG GGT CTA CAT GGC AAC TG HPRT F: TGA CAC TGG CAA AAC AAT GCA HPRT R: GGT CCT TTT CAC CAG CAA GCT FDXR (HS01031617_ml) DDB2 (Hs00172068_ml), FDXR (HS01031617_ml) RNA amount used for cDNA synthesis 0.2 μg; 1.65 μg per array Cycles 1×/50°C/2min, 95°C/10min, 4O×/95°C/15 s, 60°C/1min 1×/95°C/10min, 40×/95°C/15 s, 60°C/1min 1×/95°C/5min, 40×/95°C/10 s, 60°C/30s 1×/95°C/15min, 45×/95°C/15s, 60°C/20s, 72°C/20s, 1×/95°C/5s, 1×/65°C/60s, 1×/97°C, cooling step TaqMan ® assay: 1 ×/50°C/2min, 1×/95°C/20s, 45×/95°C/3s, 60°C/30s SYBR Green assay: 1×/95°C/10min, 40×/95°C/15s, 59°C/30s, 72°C/30s, 1×/72°C/1min 1×/95°C/2min, 40×/95°C/10s, 60°C/1min 1× /50°C/2min, 1× /95°C/10min, 40×/95°C/1min, 60°C/1min cDNA amount used for cRNA synthesis N.D.
Polymerase Chain Reaction:Article Title: RENEB Inter-Laboratory Comparison 2021: The Gene Expression Assay
Article Snippet: .. Additionally, all column RNA prep kits remove most of the DNA. (−) RT control conventional PCR (ß-actin primer, HotStar MasterMix (Qiagen), 30 cycles) Check DNA conta mination No cDNA synthesis cDNA synthesis Kit/MasterMix High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) QuantiTect Reverse Transcription (Qiagen) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) RevertAid First Strand cDNA Synthesis Kit (Thermo Scientific) High Capacity cDNA Archive Kit High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) Kit/MasterMix Quick Amp Labeling Kit (Agilent) PCR protocol 1× /25°C/10min, 1×/37°C/ 120min, 1×/85°C/5min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/42°C/2min, 1×/42°C/20min, 1×/95°C/3min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/25°C/5min, 1×/42°C/60min, 1×/l0°C/5min 1×/25°C/10min, 1×/37°C/120min, 1× /85°C/5min 1×/25°C/10min, 1×/37°C/120min PCR protocol 1×/40°C/120min, 1×/70°C/15min; 1 × /40°C/120min Quality control UBC Ct ITFG1 Ct, DPM1 Ct MRPS5 Ct No HPRT1 Ct 18S rRNA Ct Quality control NanoDrop TM qRT-PCR Kit/MasterMix TaqMan Universal Master Mix TaqMan Universal Master Mix II, no UNG (Thermo Fisher Scientific) QuantiFast SYBR Green PCR (Qiagen) 5X HOT FIREPol ® EvaGreen ® qPCR SuperMix, Solis BioDyne TaqMan fast advanced master mix (Applied Biosystems) and Maxima SYBR Green qPCR Master Mix (Thermo Scientific) TaqMan,PerfeCTa ® , MultiPlex qPCR SuperMix, Quanta bioscience TaqMan Universal Master Mix Microarray DNA-Microarray Agilent, 44k whole human genome, G4112F TaqMan assays SYBR Green assay FDXR (Hs00244586_ml), GDF15 (Hs00171132_ml) BAX (Hs00180269_ml), BBC3 (Hs00248075_ml), CDKN1A (Hs00355782_ml), DDB2 (Hs03044953_ml), FDXR (Hs00244586_ml), GADD45A (Hs00169255_ml), GDF15 (Hs00171132_ml), TNFSF4 (Hs00182411_ml) CDKN1A-F: AGACCAGCATGACAGATTTCTACC; CDKN1A-R: CTTCCTGTGGGCGGATTAGG; DDB2-F: AGCATCACTGGGCTGAAGTT; DDB2-R: TGGTGTCTGAGCTGGCAAAA; FDX-F: TGGAGAGAACGGACATCACG; FDX-R: AGCCACACTGTCTTCACTCG GADD45a for: ACTGCGTGCTGGTGACGAAT, GADD45a rev: GTTGACTTAAGGCAGGATCCTTCCA; FDXR for: TGGATGTGCCAGGCCTCTAC, FDXR rev: TGAGGAAGCTGTCAGTCATGGTT; CDKN1A for: CCTGGAGACTCTCAGGGTCGAAA, CDKN1A rev: GCGTTTGGAGTGGTAGAAATCTGTCA; MDM2 for: TATCAGGCAGGGGAGAGTGATACA, MDM2 rev: CCAACATCTGTTGCAATGTGATGGAA; 18S for: GCTTAATTTGACTCAACACGGGA, 18S rev: AGCTATCAATCTGTCAATCCTGTCC. .. TaqMan ® assay: DDB2 (Hs00172068_ml), FDXR (HS01031617_ml), ITFG1: Hs01061271_ml SYBR Green assay: CDKN1A F:CCT CAT CCC GTG TTC TCC TTT CDKN1A R: GTA CCA CCC AGC GGA CAA GT GAPDH F: CGA CCA CTT TGT CAA GCT CA GAPDH R: AGG GGT CTA CAT GGC AAC TG HPRT F: TGA CAC TGG CAA AAC AAT GCA HPRT R: GGT CCT TTT CAC CAG CAA GCT FDXR (HS01031617_ml) DDB2 (Hs00172068_ml), FDXR (HS01031617_ml) RNA amount used for cDNA synthesis 0.2 μg; 1.65 μg per array Cycles 1×/50°C/2min, 95°C/10min, 4O×/95°C/15 s, 60°C/1min 1×/95°C/10min, 40×/95°C/15 s, 60°C/1min 1×/95°C/5min, 40×/95°C/10 s, 60°C/30s 1×/95°C/15min, 45×/95°C/15s, 60°C/20s, 72°C/20s, 1×/95°C/5s, 1×/65°C/60s, 1×/97°C, cooling step TaqMan ® assay: 1 ×/50°C/2min, 1×/95°C/20s, 45×/95°C/3s, 60°C/30s SYBR Green assay: 1×/95°C/10min, 40×/95°C/15s, 59°C/30s, 72°C/30s, 1×/72°C/1min 1×/95°C/2min, 40×/95°C/10s, 60°C/1min 1× /50°C/2min, 1× /95°C/10min, 40×/95°C/1min, 60°C/1min cDNA amount used for cRNA synthesis N.D.
cDNA Synthesis:Article Title: RENEB Inter-Laboratory Comparison 2021: The Gene Expression Assay
Article Snippet: .. Additionally, all column RNA prep kits remove most of the DNA. (−) RT control conventional PCR (ß-actin primer, HotStar MasterMix (Qiagen), 30 cycles) Check DNA conta mination No cDNA synthesis cDNA synthesis Kit/MasterMix High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) QuantiTect Reverse Transcription (Qiagen) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) RevertAid First Strand cDNA Synthesis Kit (Thermo Scientific) High Capacity cDNA Archive Kit High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) Kit/MasterMix Quick Amp Labeling Kit (Agilent) PCR protocol 1× /25°C/10min, 1×/37°C/ 120min, 1×/85°C/5min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/42°C/2min, 1×/42°C/20min, 1×/95°C/3min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/25°C/5min, 1×/42°C/60min, 1×/l0°C/5min 1×/25°C/10min, 1×/37°C/120min, 1× /85°C/5min 1×/25°C/10min, 1×/37°C/120min PCR protocol 1×/40°C/120min, 1×/70°C/15min; 1 × /40°C/120min Quality control UBC Ct ITFG1 Ct, DPM1 Ct MRPS5 Ct No HPRT1 Ct 18S rRNA Ct Quality control NanoDrop TM qRT-PCR Kit/MasterMix TaqMan Universal Master Mix TaqMan Universal Master Mix II, no UNG (Thermo Fisher Scientific) QuantiFast SYBR Green PCR (Qiagen) 5X HOT FIREPol ® EvaGreen ® qPCR SuperMix, Solis BioDyne TaqMan fast advanced master mix (Applied Biosystems) and Maxima SYBR Green qPCR Master Mix (Thermo Scientific) TaqMan,PerfeCTa ® , MultiPlex qPCR SuperMix, Quanta bioscience TaqMan Universal Master Mix Microarray DNA-Microarray Agilent, 44k whole human genome, G4112F TaqMan assays SYBR Green assay FDXR (Hs00244586_ml), GDF15 (Hs00171132_ml) BAX (Hs00180269_ml), BBC3 (Hs00248075_ml), CDKN1A (Hs00355782_ml), DDB2 (Hs03044953_ml), FDXR (Hs00244586_ml), GADD45A (Hs00169255_ml), GDF15 (Hs00171132_ml), TNFSF4 (Hs00182411_ml) CDKN1A-F: AGACCAGCATGACAGATTTCTACC; CDKN1A-R: CTTCCTGTGGGCGGATTAGG; DDB2-F: AGCATCACTGGGCTGAAGTT; DDB2-R: TGGTGTCTGAGCTGGCAAAA; FDX-F: TGGAGAGAACGGACATCACG; FDX-R: AGCCACACTGTCTTCACTCG GADD45a for: ACTGCGTGCTGGTGACGAAT, GADD45a rev: GTTGACTTAAGGCAGGATCCTTCCA; FDXR for: TGGATGTGCCAGGCCTCTAC, FDXR rev: TGAGGAAGCTGTCAGTCATGGTT; CDKN1A for: CCTGGAGACTCTCAGGGTCGAAA, CDKN1A rev: GCGTTTGGAGTGGTAGAAATCTGTCA; MDM2 for: TATCAGGCAGGGGAGAGTGATACA, MDM2 rev: CCAACATCTGTTGCAATGTGATGGAA; 18S for: GCTTAATTTGACTCAACACGGGA, 18S rev: AGCTATCAATCTGTCAATCCTGTCC. .. TaqMan ® assay: DDB2 (Hs00172068_ml), FDXR (HS01031617_ml), ITFG1: Hs01061271_ml SYBR Green assay: CDKN1A F:CCT CAT CCC GTG TTC TCC TTT CDKN1A R: GTA CCA CCC AGC GGA CAA GT GAPDH F: CGA CCA CTT TGT CAA GCT CA GAPDH R: AGG GGT CTA CAT GGC AAC TG HPRT F: TGA CAC TGG CAA AAC AAT GCA HPRT R: GGT CCT TTT CAC CAG CAA GCT FDXR (HS01031617_ml) DDB2 (Hs00172068_ml), FDXR (HS01031617_ml) RNA amount used for cDNA synthesis 0.2 μg; 1.65 μg per array Cycles 1×/50°C/2min, 95°C/10min, 4O×/95°C/15 s, 60°C/1min 1×/95°C/10min, 40×/95°C/15 s, 60°C/1min 1×/95°C/5min, 40×/95°C/10 s, 60°C/30s 1×/95°C/15min, 45×/95°C/15s, 60°C/20s, 72°C/20s, 1×/95°C/5s, 1×/65°C/60s, 1×/97°C, cooling step TaqMan ® assay: 1 ×/50°C/2min, 1×/95°C/20s, 45×/95°C/3s, 60°C/30s SYBR Green assay: 1×/95°C/10min, 40×/95°C/15s, 59°C/30s, 72°C/30s, 1×/72°C/1min 1×/95°C/2min, 40×/95°C/10s, 60°C/1min 1× /50°C/2min, 1× /95°C/10min, 40×/95°C/1min, 60°C/1min cDNA amount used for cRNA synthesis N.D.
Article Title: Anti-Diabetic Effect of Lactobacillus Paracasei Isolated from Malaysian Water Kefir Grains.
Article Snippet: The prevalence of type 2 diabetes mellitus (T2DM) is alarming because it is always linked to the increase in chronic diseases, mortality, and socioeconomic burden.. Water kefir has a wide range of functional and probiotic characteristics attributed to the microorganisms present in the kefir grains.. The present study aims to evaluate the in vivo anti-diabetic potential of the isolated Lactobacillus paracasei from Malaysian water kefir grains (MWKG) which was reported to have excellent probiotic properties and high antioxidant activities as reported previously.
Reverse Transcription:Article Title: RENEB Inter-Laboratory Comparison 2021: The Gene Expression Assay
Article Snippet: .. Additionally, all column RNA prep kits remove most of the DNA. (−) RT control conventional PCR (ß-actin primer, HotStar MasterMix (Qiagen), 30 cycles) Check DNA conta mination No cDNA synthesis cDNA synthesis Kit/MasterMix High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) QuantiTect Reverse Transcription (Qiagen) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) RevertAid First Strand cDNA Synthesis Kit (Thermo Scientific) High Capacity cDNA Archive Kit High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) Kit/MasterMix Quick Amp Labeling Kit (Agilent) PCR protocol 1× /25°C/10min, 1×/37°C/ 120min, 1×/85°C/5min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/42°C/2min, 1×/42°C/20min, 1×/95°C/3min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/25°C/5min, 1×/42°C/60min, 1×/l0°C/5min 1×/25°C/10min, 1×/37°C/120min, 1× /85°C/5min 1×/25°C/10min, 1×/37°C/120min PCR protocol 1×/40°C/120min, 1×/70°C/15min; 1 × /40°C/120min Quality control UBC Ct ITFG1 Ct, DPM1 Ct MRPS5 Ct No HPRT1 Ct 18S rRNA Ct Quality control NanoDrop TM qRT-PCR Kit/MasterMix TaqMan Universal Master Mix TaqMan Universal Master Mix II, no UNG (Thermo Fisher Scientific) QuantiFast SYBR Green PCR (Qiagen) 5X HOT FIREPol ® EvaGreen ® qPCR SuperMix, Solis BioDyne TaqMan fast advanced master mix (Applied Biosystems) and Maxima SYBR Green qPCR Master Mix (Thermo Scientific) TaqMan,PerfeCTa ® , MultiPlex qPCR SuperMix, Quanta bioscience TaqMan Universal Master Mix Microarray DNA-Microarray Agilent, 44k whole human genome, G4112F TaqMan assays SYBR Green assay FDXR (Hs00244586_ml), GDF15 (Hs00171132_ml) BAX (Hs00180269_ml), BBC3 (Hs00248075_ml), CDKN1A (Hs00355782_ml), DDB2 (Hs03044953_ml), FDXR (Hs00244586_ml), GADD45A (Hs00169255_ml), GDF15 (Hs00171132_ml), TNFSF4 (Hs00182411_ml) CDKN1A-F: AGACCAGCATGACAGATTTCTACC; CDKN1A-R: CTTCCTGTGGGCGGATTAGG; DDB2-F: AGCATCACTGGGCTGAAGTT; DDB2-R: TGGTGTCTGAGCTGGCAAAA; FDX-F: TGGAGAGAACGGACATCACG; FDX-R: AGCCACACTGTCTTCACTCG GADD45a for: ACTGCGTGCTGGTGACGAAT, GADD45a rev: GTTGACTTAAGGCAGGATCCTTCCA; FDXR for: TGGATGTGCCAGGCCTCTAC, FDXR rev: TGAGGAAGCTGTCAGTCATGGTT; CDKN1A for: CCTGGAGACTCTCAGGGTCGAAA, CDKN1A rev: GCGTTTGGAGTGGTAGAAATCTGTCA; MDM2 for: TATCAGGCAGGGGAGAGTGATACA, MDM2 rev: CCAACATCTGTTGCAATGTGATGGAA; 18S for: GCTTAATTTGACTCAACACGGGA, 18S rev: AGCTATCAATCTGTCAATCCTGTCC. .. TaqMan ® assay: DDB2 (Hs00172068_ml), FDXR (HS01031617_ml), ITFG1: Hs01061271_ml SYBR Green assay: CDKN1A F:CCT CAT CCC GTG TTC TCC TTT CDKN1A R: GTA CCA CCC AGC GGA CAA GT GAPDH F: CGA CCA CTT TGT CAA GCT CA GAPDH R: AGG GGT CTA CAT GGC AAC TG HPRT F: TGA CAC TGG CAA AAC AAT GCA HPRT R: GGT CCT TTT CAC CAG CAA GCT FDXR (HS01031617_ml) DDB2 (Hs00172068_ml), FDXR (HS01031617_ml) RNA amount used for cDNA synthesis 0.2 μg; 1.65 μg per array Cycles 1×/50°C/2min, 95°C/10min, 4O×/95°C/15 s, 60°C/1min 1×/95°C/10min, 40×/95°C/15 s, 60°C/1min 1×/95°C/5min, 40×/95°C/10 s, 60°C/30s 1×/95°C/15min, 45×/95°C/15s, 60°C/20s, 72°C/20s, 1×/95°C/5s, 1×/65°C/60s, 1×/97°C, cooling step TaqMan ® assay: 1 ×/50°C/2min, 1×/95°C/20s, 45×/95°C/3s, 60°C/30s SYBR Green assay: 1×/95°C/10min, 40×/95°C/15s, 59°C/30s, 72°C/30s, 1×/72°C/1min 1×/95°C/2min, 40×/95°C/10s, 60°C/1min 1× /50°C/2min, 1× /95°C/10min, 40×/95°C/1min, 60°C/1min cDNA amount used for cRNA synthesis N.D.
Article Title: Circ_005077 accelerates myocardial lipotoxicity induced by high-fat diet via CyPA/p47PHOX mediated ferroptosis
Article Snippet: The captured mRNA was fragmented into small pieces at 94 °C for 5–7 min using the magnesium RNA fragmentation module (E6150, NEB, USA). .. SuperScriptTM II synthesized cDNA reverse transcriptase (1,896,649, Invitrogen, USA), then E. coli DNA polymerase I (m0209, NEB, USA) and RNase H (m0297, NEB, USA) were used for double-stranded synthesis to synthesize U-labeled second-stranded DNAs, dUTP solution (R0133, Thermo Fisher, USA) was added to make the ends of double-stranded DNA flat. ..
Article Title: Anti-Diabetic Effect of Lactobacillus Paracasei Isolated from Malaysian Water Kefir Grains.
Article Snippet: The prevalence of type 2 diabetes mellitus (T2DM) is alarming because it is always linked to the increase in chronic diseases, mortality, and socioeconomic burden.. Water kefir has a wide range of functional and probiotic characteristics attributed to the microorganisms present in the kefir grains.. The present study aims to evaluate the in vivo anti-diabetic potential of the isolated Lactobacillus paracasei from Malaysian water kefir grains (MWKG) which was reported to have excellent probiotic properties and high antioxidant activities as reported previously.
Labeling:Article Title: RENEB Inter-Laboratory Comparison 2021: The Gene Expression Assay
Article Snippet: .. Additionally, all column RNA prep kits remove most of the DNA. (−) RT control conventional PCR (ß-actin primer, HotStar MasterMix (Qiagen), 30 cycles) Check DNA conta mination No cDNA synthesis cDNA synthesis Kit/MasterMix High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) QuantiTect Reverse Transcription (Qiagen) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) RevertAid First Strand cDNA Synthesis Kit (Thermo Scientific) High Capacity cDNA Archive Kit High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) Kit/MasterMix Quick Amp Labeling Kit (Agilent) PCR protocol 1× /25°C/10min, 1×/37°C/ 120min, 1×/85°C/5min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/42°C/2min, 1×/42°C/20min, 1×/95°C/3min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/25°C/5min, 1×/42°C/60min, 1×/l0°C/5min 1×/25°C/10min, 1×/37°C/120min, 1× /85°C/5min 1×/25°C/10min, 1×/37°C/120min PCR protocol 1×/40°C/120min, 1×/70°C/15min; 1 × /40°C/120min Quality control UBC Ct ITFG1 Ct, DPM1 Ct MRPS5 Ct No HPRT1 Ct 18S rRNA Ct Quality control NanoDrop TM qRT-PCR Kit/MasterMix TaqMan Universal Master Mix TaqMan Universal Master Mix II, no UNG (Thermo Fisher Scientific) QuantiFast SYBR Green PCR (Qiagen) 5X HOT FIREPol ® EvaGreen ® qPCR SuperMix, Solis BioDyne TaqMan fast advanced master mix (Applied Biosystems) and Maxima SYBR Green qPCR Master Mix (Thermo Scientific) TaqMan,PerfeCTa ® , MultiPlex qPCR SuperMix, Quanta bioscience TaqMan Universal Master Mix Microarray DNA-Microarray Agilent, 44k whole human genome, G4112F TaqMan assays SYBR Green assay FDXR (Hs00244586_ml), GDF15 (Hs00171132_ml) BAX (Hs00180269_ml), BBC3 (Hs00248075_ml), CDKN1A (Hs00355782_ml), DDB2 (Hs03044953_ml), FDXR (Hs00244586_ml), GADD45A (Hs00169255_ml), GDF15 (Hs00171132_ml), TNFSF4 (Hs00182411_ml) CDKN1A-F: AGACCAGCATGACAGATTTCTACC; CDKN1A-R: CTTCCTGTGGGCGGATTAGG; DDB2-F: AGCATCACTGGGCTGAAGTT; DDB2-R: TGGTGTCTGAGCTGGCAAAA; FDX-F: TGGAGAGAACGGACATCACG; FDX-R: AGCCACACTGTCTTCACTCG GADD45a for: ACTGCGTGCTGGTGACGAAT, GADD45a rev: GTTGACTTAAGGCAGGATCCTTCCA; FDXR for: TGGATGTGCCAGGCCTCTAC, FDXR rev: TGAGGAAGCTGTCAGTCATGGTT; CDKN1A for: CCTGGAGACTCTCAGGGTCGAAA, CDKN1A rev: GCGTTTGGAGTGGTAGAAATCTGTCA; MDM2 for: TATCAGGCAGGGGAGAGTGATACA, MDM2 rev: CCAACATCTGTTGCAATGTGATGGAA; 18S for: GCTTAATTTGACTCAACACGGGA, 18S rev: AGCTATCAATCTGTCAATCCTGTCC. .. TaqMan ® assay: DDB2 (Hs00172068_ml), FDXR (HS01031617_ml), ITFG1: Hs01061271_ml SYBR Green assay: CDKN1A F:CCT CAT CCC GTG TTC TCC TTT CDKN1A R: GTA CCA CCC AGC GGA CAA GT GAPDH F: CGA CCA CTT TGT CAA GCT CA GAPDH R: AGG GGT CTA CAT GGC AAC TG HPRT F: TGA CAC TGG CAA AAC AAT GCA HPRT R: GGT CCT TTT CAC CAG CAA GCT FDXR (HS01031617_ml) DDB2 (Hs00172068_ml), FDXR (HS01031617_ml) RNA amount used for cDNA synthesis 0.2 μg; 1.65 μg per array Cycles 1×/50°C/2min, 95°C/10min, 4O×/95°C/15 s, 60°C/1min 1×/95°C/10min, 40×/95°C/15 s, 60°C/1min 1×/95°C/5min, 40×/95°C/10 s, 60°C/30s 1×/95°C/15min, 45×/95°C/15s, 60°C/20s, 72°C/20s, 1×/95°C/5s, 1×/65°C/60s, 1×/97°C, cooling step TaqMan ® assay: 1 ×/50°C/2min, 1×/95°C/20s, 45×/95°C/3s, 60°C/30s SYBR Green assay: 1×/95°C/10min, 40×/95°C/15s, 59°C/30s, 72°C/30s, 1×/72°C/1min 1×/95°C/2min, 40×/95°C/10s, 60°C/1min 1× /50°C/2min, 1× /95°C/10min, 40×/95°C/1min, 60°C/1min cDNA amount used for cRNA synthesis N.D.
Quantitative RT-PCR:Article Title: RENEB Inter-Laboratory Comparison 2021: The Gene Expression Assay
Article Snippet: .. Additionally, all column RNA prep kits remove most of the DNA. (−) RT control conventional PCR (ß-actin primer, HotStar MasterMix (Qiagen), 30 cycles) Check DNA conta mination No cDNA synthesis cDNA synthesis Kit/MasterMix High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) QuantiTect Reverse Transcription (Qiagen) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) RevertAid First Strand cDNA Synthesis Kit (Thermo Scientific) High Capacity cDNA Archive Kit High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) Kit/MasterMix Quick Amp Labeling Kit (Agilent) PCR protocol 1× /25°C/10min, 1×/37°C/ 120min, 1×/85°C/5min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/42°C/2min, 1×/42°C/20min, 1×/95°C/3min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/25°C/5min, 1×/42°C/60min, 1×/l0°C/5min 1×/25°C/10min, 1×/37°C/120min, 1× /85°C/5min 1×/25°C/10min, 1×/37°C/120min PCR protocol 1×/40°C/120min, 1×/70°C/15min; 1 × /40°C/120min Quality control UBC Ct ITFG1 Ct, DPM1 Ct MRPS5 Ct No HPRT1 Ct 18S rRNA Ct Quality control NanoDrop TM qRT-PCR Kit/MasterMix TaqMan Universal Master Mix TaqMan Universal Master Mix II, no UNG (Thermo Fisher Scientific) QuantiFast SYBR Green PCR (Qiagen) 5X HOT FIREPol ® EvaGreen ® qPCR SuperMix, Solis BioDyne TaqMan fast advanced master mix (Applied Biosystems) and Maxima SYBR Green qPCR Master Mix (Thermo Scientific) TaqMan,PerfeCTa ® , MultiPlex qPCR SuperMix, Quanta bioscience TaqMan Universal Master Mix Microarray DNA-Microarray Agilent, 44k whole human genome, G4112F TaqMan assays SYBR Green assay FDXR (Hs00244586_ml), GDF15 (Hs00171132_ml) BAX (Hs00180269_ml), BBC3 (Hs00248075_ml), CDKN1A (Hs00355782_ml), DDB2 (Hs03044953_ml), FDXR (Hs00244586_ml), GADD45A (Hs00169255_ml), GDF15 (Hs00171132_ml), TNFSF4 (Hs00182411_ml) CDKN1A-F: AGACCAGCATGACAGATTTCTACC; CDKN1A-R: CTTCCTGTGGGCGGATTAGG; DDB2-F: AGCATCACTGGGCTGAAGTT; DDB2-R: TGGTGTCTGAGCTGGCAAAA; FDX-F: TGGAGAGAACGGACATCACG; FDX-R: AGCCACACTGTCTTCACTCG GADD45a for: ACTGCGTGCTGGTGACGAAT, GADD45a rev: GTTGACTTAAGGCAGGATCCTTCCA; FDXR for: TGGATGTGCCAGGCCTCTAC, FDXR rev: TGAGGAAGCTGTCAGTCATGGTT; CDKN1A for: CCTGGAGACTCTCAGGGTCGAAA, CDKN1A rev: GCGTTTGGAGTGGTAGAAATCTGTCA; MDM2 for: TATCAGGCAGGGGAGAGTGATACA, MDM2 rev: CCAACATCTGTTGCAATGTGATGGAA; 18S for: GCTTAATTTGACTCAACACGGGA, 18S rev: AGCTATCAATCTGTCAATCCTGTCC. .. TaqMan ® assay: DDB2 (Hs00172068_ml), FDXR (HS01031617_ml), ITFG1: Hs01061271_ml SYBR Green assay: CDKN1A F:CCT CAT CCC GTG TTC TCC TTT CDKN1A R: GTA CCA CCC AGC GGA CAA GT GAPDH F: CGA CCA CTT TGT CAA GCT CA GAPDH R: AGG GGT CTA CAT GGC AAC TG HPRT F: TGA CAC TGG CAA AAC AAT GCA HPRT R: GGT CCT TTT CAC CAG CAA GCT FDXR (HS01031617_ml) DDB2 (Hs00172068_ml), FDXR (HS01031617_ml) RNA amount used for cDNA synthesis 0.2 μg; 1.65 μg per array Cycles 1×/50°C/2min, 95°C/10min, 4O×/95°C/15 s, 60°C/1min 1×/95°C/10min, 40×/95°C/15 s, 60°C/1min 1×/95°C/5min, 40×/95°C/10 s, 60°C/30s 1×/95°C/15min, 45×/95°C/15s, 60°C/20s, 72°C/20s, 1×/95°C/5s, 1×/65°C/60s, 1×/97°C, cooling step TaqMan ® assay: 1 ×/50°C/2min, 1×/95°C/20s, 45×/95°C/3s, 60°C/30s SYBR Green assay: 1×/95°C/10min, 40×/95°C/15s, 59°C/30s, 72°C/30s, 1×/72°C/1min 1×/95°C/2min, 40×/95°C/10s, 60°C/1min 1× /50°C/2min, 1× /95°C/10min, 40×/95°C/1min, 60°C/1min cDNA amount used for cRNA synthesis N.D.
SYBR Green Assay:Article Title: RENEB Inter-Laboratory Comparison 2021: The Gene Expression Assay
Article Snippet: .. Additionally, all column RNA prep kits remove most of the DNA. (−) RT control conventional PCR (ß-actin primer, HotStar MasterMix (Qiagen), 30 cycles) Check DNA conta mination No cDNA synthesis cDNA synthesis Kit/MasterMix High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) QuantiTect Reverse Transcription (Qiagen) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) RevertAid First Strand cDNA Synthesis Kit (Thermo Scientific) High Capacity cDNA Archive Kit High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) Kit/MasterMix Quick Amp Labeling Kit (Agilent) PCR protocol 1× /25°C/10min, 1×/37°C/ 120min, 1×/85°C/5min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/42°C/2min, 1×/42°C/20min, 1×/95°C/3min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/25°C/5min, 1×/42°C/60min, 1×/l0°C/5min 1×/25°C/10min, 1×/37°C/120min, 1× /85°C/5min 1×/25°C/10min, 1×/37°C/120min PCR protocol 1×/40°C/120min, 1×/70°C/15min; 1 × /40°C/120min Quality control UBC Ct ITFG1 Ct, DPM1 Ct MRPS5 Ct No HPRT1 Ct 18S rRNA Ct Quality control NanoDrop TM qRT-PCR Kit/MasterMix TaqMan Universal Master Mix TaqMan Universal Master Mix II, no UNG (Thermo Fisher Scientific) QuantiFast SYBR Green PCR (Qiagen) 5X HOT FIREPol ® EvaGreen ® qPCR SuperMix, Solis BioDyne TaqMan fast advanced master mix (Applied Biosystems) and Maxima SYBR Green qPCR Master Mix (Thermo Scientific) TaqMan,PerfeCTa ® , MultiPlex qPCR SuperMix, Quanta bioscience TaqMan Universal Master Mix Microarray DNA-Microarray Agilent, 44k whole human genome, G4112F TaqMan assays SYBR Green assay FDXR (Hs00244586_ml), GDF15 (Hs00171132_ml) BAX (Hs00180269_ml), BBC3 (Hs00248075_ml), CDKN1A (Hs00355782_ml), DDB2 (Hs03044953_ml), FDXR (Hs00244586_ml), GADD45A (Hs00169255_ml), GDF15 (Hs00171132_ml), TNFSF4 (Hs00182411_ml) CDKN1A-F: AGACCAGCATGACAGATTTCTACC; CDKN1A-R: CTTCCTGTGGGCGGATTAGG; DDB2-F: AGCATCACTGGGCTGAAGTT; DDB2-R: TGGTGTCTGAGCTGGCAAAA; FDX-F: TGGAGAGAACGGACATCACG; FDX-R: AGCCACACTGTCTTCACTCG GADD45a for: ACTGCGTGCTGGTGACGAAT, GADD45a rev: GTTGACTTAAGGCAGGATCCTTCCA; FDXR for: TGGATGTGCCAGGCCTCTAC, FDXR rev: TGAGGAAGCTGTCAGTCATGGTT; CDKN1A for: CCTGGAGACTCTCAGGGTCGAAA, CDKN1A rev: GCGTTTGGAGTGGTAGAAATCTGTCA; MDM2 for: TATCAGGCAGGGGAGAGTGATACA, MDM2 rev: CCAACATCTGTTGCAATGTGATGGAA; 18S for: GCTTAATTTGACTCAACACGGGA, 18S rev: AGCTATCAATCTGTCAATCCTGTCC. .. TaqMan ® assay: DDB2 (Hs00172068_ml), FDXR (HS01031617_ml), ITFG1: Hs01061271_ml SYBR Green assay: CDKN1A F:CCT CAT CCC GTG TTC TCC TTT CDKN1A R: GTA CCA CCC AGC GGA CAA GT GAPDH F: CGA CCA CTT TGT CAA GCT CA GAPDH R: AGG GGT CTA CAT GGC AAC TG HPRT F: TGA CAC TGG CAA AAC AAT GCA HPRT R: GGT CCT TTT CAC CAG CAA GCT FDXR (HS01031617_ml) DDB2 (Hs00172068_ml), FDXR (HS01031617_ml) RNA amount used for cDNA synthesis 0.2 μg; 1.65 μg per array Cycles 1×/50°C/2min, 95°C/10min, 4O×/95°C/15 s, 60°C/1min 1×/95°C/10min, 40×/95°C/15 s, 60°C/1min 1×/95°C/5min, 40×/95°C/10 s, 60°C/30s 1×/95°C/15min, 45×/95°C/15s, 60°C/20s, 72°C/20s, 1×/95°C/5s, 1×/65°C/60s, 1×/97°C, cooling step TaqMan ® assay: 1 ×/50°C/2min, 1×/95°C/20s, 45×/95°C/3s, 60°C/30s SYBR Green assay: 1×/95°C/10min, 40×/95°C/15s, 59°C/30s, 72°C/30s, 1×/72°C/1min 1×/95°C/2min, 40×/95°C/10s, 60°C/1min 1× /50°C/2min, 1× /95°C/10min, 40×/95°C/1min, 60°C/1min cDNA amount used for cRNA synthesis N.D.
Article Title: Anti-Diabetic Effect of Lactobacillus Paracasei Isolated from Malaysian Water Kefir Grains.
Article Snippet: The prevalence of type 2 diabetes mellitus (T2DM) is alarming because it is always linked to the increase in chronic diseases, mortality, and socioeconomic burden.. Water kefir has a wide range of functional and probiotic characteristics attributed to the microorganisms present in the kefir grains.. The present study aims to evaluate the in vivo anti-diabetic potential of the isolated Lactobacillus paracasei from Malaysian water kefir grains (MWKG) which was reported to have excellent probiotic properties and high antioxidant activities as reported previously.
Real-time Polymerase Chain Reaction:Article Title: RENEB Inter-Laboratory Comparison 2021: The Gene Expression Assay
Article Snippet: .. Additionally, all column RNA prep kits remove most of the DNA. (−) RT control conventional PCR (ß-actin primer, HotStar MasterMix (Qiagen), 30 cycles) Check DNA conta mination No cDNA synthesis cDNA synthesis Kit/MasterMix High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) QuantiTect Reverse Transcription (Qiagen) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) RevertAid First Strand cDNA Synthesis Kit (Thermo Scientific) High Capacity cDNA Archive Kit High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) Kit/MasterMix Quick Amp Labeling Kit (Agilent) PCR protocol 1× /25°C/10min, 1×/37°C/ 120min, 1×/85°C/5min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/42°C/2min, 1×/42°C/20min, 1×/95°C/3min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/25°C/5min, 1×/42°C/60min, 1×/l0°C/5min 1×/25°C/10min, 1×/37°C/120min, 1× /85°C/5min 1×/25°C/10min, 1×/37°C/120min PCR protocol 1×/40°C/120min, 1×/70°C/15min; 1 × /40°C/120min Quality control UBC Ct ITFG1 Ct, DPM1 Ct MRPS5 Ct No HPRT1 Ct 18S rRNA Ct Quality control NanoDrop TM qRT-PCR Kit/MasterMix TaqMan Universal Master Mix TaqMan Universal Master Mix II, no UNG (Thermo Fisher Scientific) QuantiFast SYBR Green PCR (Qiagen) 5X HOT FIREPol ® EvaGreen ® qPCR SuperMix, Solis BioDyne TaqMan fast advanced master mix (Applied Biosystems) and Maxima SYBR Green qPCR Master Mix (Thermo Scientific) TaqMan,PerfeCTa ® , MultiPlex qPCR SuperMix, Quanta bioscience TaqMan Universal Master Mix Microarray DNA-Microarray Agilent, 44k whole human genome, G4112F TaqMan assays SYBR Green assay FDXR (Hs00244586_ml), GDF15 (Hs00171132_ml) BAX (Hs00180269_ml), BBC3 (Hs00248075_ml), CDKN1A (Hs00355782_ml), DDB2 (Hs03044953_ml), FDXR (Hs00244586_ml), GADD45A (Hs00169255_ml), GDF15 (Hs00171132_ml), TNFSF4 (Hs00182411_ml) CDKN1A-F: AGACCAGCATGACAGATTTCTACC; CDKN1A-R: CTTCCTGTGGGCGGATTAGG; DDB2-F: AGCATCACTGGGCTGAAGTT; DDB2-R: TGGTGTCTGAGCTGGCAAAA; FDX-F: TGGAGAGAACGGACATCACG; FDX-R: AGCCACACTGTCTTCACTCG GADD45a for: ACTGCGTGCTGGTGACGAAT, GADD45a rev: GTTGACTTAAGGCAGGATCCTTCCA; FDXR for: TGGATGTGCCAGGCCTCTAC, FDXR rev: TGAGGAAGCTGTCAGTCATGGTT; CDKN1A for: CCTGGAGACTCTCAGGGTCGAAA, CDKN1A rev: GCGTTTGGAGTGGTAGAAATCTGTCA; MDM2 for: TATCAGGCAGGGGAGAGTGATACA, MDM2 rev: CCAACATCTGTTGCAATGTGATGGAA; 18S for: GCTTAATTTGACTCAACACGGGA, 18S rev: AGCTATCAATCTGTCAATCCTGTCC. .. TaqMan ® assay: DDB2 (Hs00172068_ml), FDXR (HS01031617_ml), ITFG1: Hs01061271_ml SYBR Green assay: CDKN1A F:CCT CAT CCC GTG TTC TCC TTT CDKN1A R: GTA CCA CCC AGC GGA CAA GT GAPDH F: CGA CCA CTT TGT CAA GCT CA GAPDH R: AGG GGT CTA CAT GGC AAC TG HPRT F: TGA CAC TGG CAA AAC AAT GCA HPRT R: GGT CCT TTT CAC CAG CAA GCT FDXR (HS01031617_ml) DDB2 (Hs00172068_ml), FDXR (HS01031617_ml) RNA amount used for cDNA synthesis 0.2 μg; 1.65 μg per array Cycles 1×/50°C/2min, 95°C/10min, 4O×/95°C/15 s, 60°C/1min 1×/95°C/10min, 40×/95°C/15 s, 60°C/1min 1×/95°C/5min, 40×/95°C/10 s, 60°C/30s 1×/95°C/15min, 45×/95°C/15s, 60°C/20s, 72°C/20s, 1×/95°C/5s, 1×/65°C/60s, 1×/97°C, cooling step TaqMan ® assay: 1 ×/50°C/2min, 1×/95°C/20s, 45×/95°C/3s, 60°C/30s SYBR Green assay: 1×/95°C/10min, 40×/95°C/15s, 59°C/30s, 72°C/30s, 1×/72°C/1min 1×/95°C/2min, 40×/95°C/10s, 60°C/1min 1× /50°C/2min, 1× /95°C/10min, 40×/95°C/1min, 60°C/1min cDNA amount used for cRNA synthesis N.D.
Article Title: Anti-Diabetic Effect of Lactobacillus Paracasei Isolated from Malaysian Water Kefir Grains.
Article Snippet: The prevalence of type 2 diabetes mellitus (T2DM) is alarming because it is always linked to the increase in chronic diseases, mortality, and socioeconomic burden.. Water kefir has a wide range of functional and probiotic characteristics attributed to the microorganisms present in the kefir grains.. The present study aims to evaluate the in vivo anti-diabetic potential of the isolated Lactobacillus paracasei from Malaysian water kefir grains (MWKG) which was reported to have excellent probiotic properties and high antioxidant activities as reported previously.
Multiplex Assay:Article Title: RENEB Inter-Laboratory Comparison 2021: The Gene Expression Assay
Article Snippet: .. Additionally, all column RNA prep kits remove most of the DNA. (−) RT control conventional PCR (ß-actin primer, HotStar MasterMix (Qiagen), 30 cycles) Check DNA conta mination No cDNA synthesis cDNA synthesis Kit/MasterMix High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) QuantiTect Reverse Transcription (Qiagen) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) RevertAid First Strand cDNA Synthesis Kit (Thermo Scientific) High Capacity cDNA Archive Kit High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) Kit/MasterMix Quick Amp Labeling Kit (Agilent) PCR protocol 1× /25°C/10min, 1×/37°C/ 120min, 1×/85°C/5min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/42°C/2min, 1×/42°C/20min, 1×/95°C/3min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/25°C/5min, 1×/42°C/60min, 1×/l0°C/5min 1×/25°C/10min, 1×/37°C/120min, 1× /85°C/5min 1×/25°C/10min, 1×/37°C/120min PCR protocol 1×/40°C/120min, 1×/70°C/15min; 1 × /40°C/120min Quality control UBC Ct ITFG1 Ct, DPM1 Ct MRPS5 Ct No HPRT1 Ct 18S rRNA Ct Quality control NanoDrop TM qRT-PCR Kit/MasterMix TaqMan Universal Master Mix TaqMan Universal Master Mix II, no UNG (Thermo Fisher Scientific) QuantiFast SYBR Green PCR (Qiagen) 5X HOT FIREPol ® EvaGreen ® qPCR SuperMix, Solis BioDyne TaqMan fast advanced master mix (Applied Biosystems) and Maxima SYBR Green qPCR Master Mix (Thermo Scientific) TaqMan,PerfeCTa ® , MultiPlex qPCR SuperMix, Quanta bioscience TaqMan Universal Master Mix Microarray DNA-Microarray Agilent, 44k whole human genome, G4112F TaqMan assays SYBR Green assay FDXR (Hs00244586_ml), GDF15 (Hs00171132_ml) BAX (Hs00180269_ml), BBC3 (Hs00248075_ml), CDKN1A (Hs00355782_ml), DDB2 (Hs03044953_ml), FDXR (Hs00244586_ml), GADD45A (Hs00169255_ml), GDF15 (Hs00171132_ml), TNFSF4 (Hs00182411_ml) CDKN1A-F: AGACCAGCATGACAGATTTCTACC; CDKN1A-R: CTTCCTGTGGGCGGATTAGG; DDB2-F: AGCATCACTGGGCTGAAGTT; DDB2-R: TGGTGTCTGAGCTGGCAAAA; FDX-F: TGGAGAGAACGGACATCACG; FDX-R: AGCCACACTGTCTTCACTCG GADD45a for: ACTGCGTGCTGGTGACGAAT, GADD45a rev: GTTGACTTAAGGCAGGATCCTTCCA; FDXR for: TGGATGTGCCAGGCCTCTAC, FDXR rev: TGAGGAAGCTGTCAGTCATGGTT; CDKN1A for: CCTGGAGACTCTCAGGGTCGAAA, CDKN1A rev: GCGTTTGGAGTGGTAGAAATCTGTCA; MDM2 for: TATCAGGCAGGGGAGAGTGATACA, MDM2 rev: CCAACATCTGTTGCAATGTGATGGAA; 18S for: GCTTAATTTGACTCAACACGGGA, 18S rev: AGCTATCAATCTGTCAATCCTGTCC. .. TaqMan ® assay: DDB2 (Hs00172068_ml), FDXR (HS01031617_ml), ITFG1: Hs01061271_ml SYBR Green assay: CDKN1A F:CCT CAT CCC GTG TTC TCC TTT CDKN1A R: GTA CCA CCC AGC GGA CAA GT GAPDH F: CGA CCA CTT TGT CAA GCT CA GAPDH R: AGG GGT CTA CAT GGC AAC TG HPRT F: TGA CAC TGG CAA AAC AAT GCA HPRT R: GGT CCT TTT CAC CAG CAA GCT FDXR (HS01031617_ml) DDB2 (Hs00172068_ml), FDXR (HS01031617_ml) RNA amount used for cDNA synthesis 0.2 μg; 1.65 μg per array Cycles 1×/50°C/2min, 95°C/10min, 4O×/95°C/15 s, 60°C/1min 1×/95°C/10min, 40×/95°C/15 s, 60°C/1min 1×/95°C/5min, 40×/95°C/10 s, 60°C/30s 1×/95°C/15min, 45×/95°C/15s, 60°C/20s, 72°C/20s, 1×/95°C/5s, 1×/65°C/60s, 1×/97°C, cooling step TaqMan ® assay: 1 ×/50°C/2min, 1×/95°C/20s, 45×/95°C/3s, 60°C/30s SYBR Green assay: 1×/95°C/10min, 40×/95°C/15s, 59°C/30s, 72°C/30s, 1×/72°C/1min 1×/95°C/2min, 40×/95°C/10s, 60°C/1min 1× /50°C/2min, 1× /95°C/10min, 40×/95°C/1min, 60°C/1min cDNA amount used for cRNA synthesis N.D.
Microarray:Article Title: RENEB Inter-Laboratory Comparison 2021: The Gene Expression Assay
Article Snippet: .. Additionally, all column RNA prep kits remove most of the DNA. (−) RT control conventional PCR (ß-actin primer, HotStar MasterMix (Qiagen), 30 cycles) Check DNA conta mination No cDNA synthesis cDNA synthesis Kit/MasterMix High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) QuantiTect Reverse Transcription (Qiagen) High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) RevertAid First Strand cDNA Synthesis Kit (Thermo Scientific) High Capacity cDNA Archive Kit High Capacity cDNA Reverse Transcription Kit (Thermo Fisher Scientific) Kit/MasterMix Quick Amp Labeling Kit (Agilent) PCR protocol 1× /25°C/10min, 1×/37°C/ 120min, 1×/85°C/5min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/42°C/2min, 1×/42°C/20min, 1×/95°C/3min 1×/25°C/10min, 1×/37°C/120min, 1×/85°C/5min 1×/25°C/5min, 1×/42°C/60min, 1×/l0°C/5min 1×/25°C/10min, 1×/37°C/120min, 1× /85°C/5min 1×/25°C/10min, 1×/37°C/120min PCR protocol 1×/40°C/120min, 1×/70°C/15min; 1 × /40°C/120min Quality control UBC Ct ITFG1 Ct, DPM1 Ct MRPS5 Ct No HPRT1 Ct 18S rRNA Ct Quality control NanoDrop TM qRT-PCR Kit/MasterMix TaqMan Universal Master Mix TaqMan Universal Master Mix II, no UNG (Thermo Fisher Scientific) QuantiFast SYBR Green PCR (Qiagen) 5X HOT FIREPol ® EvaGreen ® qPCR SuperMix, Solis BioDyne TaqMan fast advanced master mix (Applied Biosystems) and Maxima SYBR Green qPCR Master Mix (Thermo Scientific) TaqMan,PerfeCTa ® , MultiPlex qPCR SuperMix, Quanta bioscience TaqMan Universal Master Mix Microarray DNA-Microarray Agilent, 44k whole human genome, G4112F TaqMan assays SYBR Green assay FDXR (Hs00244586_ml), GDF15 (Hs00171132_ml) BAX (Hs00180269_ml), BBC3 (Hs00248075_ml), CDKN1A (Hs00355782_ml), DDB2 (Hs03044953_ml), FDXR (Hs00244586_ml), GADD45A (Hs00169255_ml), GDF15 (Hs00171132_ml), TNFSF4 (Hs00182411_ml) CDKN1A-F: AGACCAGCATGACAGATTTCTACC; CDKN1A-R: CTTCCTGTGGGCGGATTAGG; DDB2-F: AGCATCACTGGGCTGAAGTT; DDB2-R: TGGTGTCTGAGCTGGCAAAA; FDX-F: TGGAGAGAACGGACATCACG; FDX-R: AGCCACACTGTCTTCACTCG GADD45a for: ACTGCGTGCTGGTGACGAAT, GADD45a rev: GTTGACTTAAGGCAGGATCCTTCCA; FDXR for: TGGATGTGCCAGGCCTCTAC, FDXR rev: TGAGGAAGCTGTCAGTCATGGTT; CDKN1A for: CCTGGAGACTCTCAGGGTCGAAA, CDKN1A rev: GCGTTTGGAGTGGTAGAAATCTGTCA; MDM2 for: TATCAGGCAGGGGAGAGTGATACA, MDM2 rev: CCAACATCTGTTGCAATGTGATGGAA; 18S for: GCTTAATTTGACTCAACACGGGA, 18S rev: AGCTATCAATCTGTCAATCCTGTCC. .. TaqMan ® assay: DDB2 (Hs00172068_ml), FDXR (HS01031617_ml), ITFG1: Hs01061271_ml SYBR Green assay: CDKN1A F:CCT CAT CCC GTG TTC TCC TTT CDKN1A R: GTA CCA CCC AGC GGA CAA GT GAPDH F: CGA CCA CTT TGT CAA GCT CA GAPDH R: AGG GGT CTA CAT GGC AAC TG HPRT F: TGA CAC TGG CAA AAC AAT GCA HPRT R: GGT CCT TTT CAC CAG CAA GCT FDXR (HS01031617_ml) DDB2 (Hs00172068_ml), FDXR (HS01031617_ml) RNA amount used for cDNA synthesis 0.2 μg; 1.65 μg per array Cycles 1×/50°C/2min, 95°C/10min, 4O×/95°C/15 s, 60°C/1min 1×/95°C/10min, 40×/95°C/15 s, 60°C/1min 1×/95°C/5min, 40×/95°C/10 s, 60°C/30s 1×/95°C/15min, 45×/95°C/15s, 60°C/20s, 72°C/20s, 1×/95°C/5s, 1×/65°C/60s, 1×/97°C, cooling step TaqMan ® assay: 1 ×/50°C/2min, 1×/95°C/20s, 45×/95°C/3s, 60°C/30s SYBR Green assay: 1×/95°C/10min, 40×/95°C/15s, 59°C/30s, 72°C/30s, 1×/72°C/1min 1×/95°C/2min, 40×/95°C/10s, 60°C/1min 1× /50°C/2min, 1× /95°C/10min, 40×/95°C/1min, 60°C/1min cDNA amount used for cRNA synthesis N.D.
Article Title: The RNA-binding protein KSRP aggravates malignant progression of clear cell renal cell carcinoma through transcriptional inhibition and post-transcriptional destabilization of the NEDD4L ubiquitin ligase.
Article Snippet: .. Numbers of stained cells were counted in at least three random microscopic fields (× 100). cDNA microarray and data analysis Transcriptional profiling data were collected from a microarray analysis of Caki-1/shCtrl and Caki-1/shKSRP cells. cDNA hybridization was performed with Affymetrix human U133 2.0 plus arrays (ThermoFisher Scientific). .. Gene expression levels were normalized as log2 values using GeneSpring software (Agilent Technologies, Santa Clara, CA, USA), and the obtained list contained genes that were up- or downregulated with greater than twofold changes in response to KSRP-KD. miRNA OneArray® and data analysis To identify which miRNAs are regulated by KSRP, a highthroughput and specific miRNA microarray (human miRNA OneArray® miRNA profiling chip) using Caki-1 cells with/without KSRP-KD was conducted by the Phalanx Biotech Group (Hsinchu, Taiwan).
Article Title: Anti-Diabetic Effect of Lactobacillus Paracasei Isolated from Malaysian Water Kefir Grains.
Article Snippet: The prevalence of type 2 diabetes mellitus (T2DM) is alarming because it is always linked to the increase in chronic diseases, mortality, and socioeconomic burden.. Water kefir has a wide range of functional and probiotic characteristics attributed to the microorganisms present in the kefir grains.. The present study aims to evaluate the in vivo anti-diabetic potential of the isolated Lactobacillus paracasei from Malaysian water kefir grains (MWKG) which was reported to have excellent probiotic properties and high antioxidant activities as reported previously.
Synthesized:Article Title: Circ_005077 accelerates myocardial lipotoxicity induced by high-fat diet via CyPA/p47PHOX mediated ferroptosis
Article Snippet: The captured mRNA was fragmented into small pieces at 94 °C for 5–7 min using the magnesium RNA fragmentation module (E6150, NEB, USA). .. SuperScriptTM II synthesized cDNA reverse transcriptase (1,896,649, Invitrogen, USA), then E. coli DNA polymerase I (m0209, NEB, USA) and RNase H (m0297, NEB, USA) were used for double-stranded synthesis to synthesize U-labeled second-stranded DNAs, dUTP solution (R0133, Thermo Fisher, USA) was added to make the ends of double-stranded DNA flat. ..
Staining:Article Title: The RNA-binding protein KSRP aggravates malignant progression of clear cell renal cell carcinoma through transcriptional inhibition and post-transcriptional destabilization of the NEDD4L ubiquitin ligase.
Article Snippet: .. Numbers of stained cells were counted in at least three random microscopic fields (× 100). cDNA microarray and data analysis Transcriptional profiling data were collected from a microarray analysis of Caki-1/shCtrl and Caki-1/shKSRP cells. cDNA hybridization was performed with Affymetrix human U133 2.0 plus arrays (ThermoFisher Scientific). .. Gene expression levels were normalized as log2 values using GeneSpring software (Agilent Technologies, Santa Clara, CA, USA), and the obtained list contained genes that were up- or downregulated with greater than twofold changes in response to KSRP-KD. miRNA OneArray® and data analysis To identify which miRNAs are regulated by KSRP, a highthroughput and specific miRNA microarray (human miRNA OneArray® miRNA profiling chip) using Caki-1 cells with/without KSRP-KD was conducted by the Phalanx Biotech Group (Hsinchu, Taiwan).
Hybridization:Article Title: The RNA-binding protein KSRP aggravates malignant progression of clear cell renal cell carcinoma through transcriptional inhibition and post-transcriptional destabilization of the NEDD4L ubiquitin ligase.
Article Snippet: .. Numbers of stained cells were counted in at least three random microscopic fields (× 100). cDNA microarray and data analysis Transcriptional profiling data were collected from a microarray analysis of Caki-1/shCtrl and Caki-1/shKSRP cells. cDNA hybridization was performed with Affymetrix human U133 2.0 plus arrays (ThermoFisher Scientific). .. Gene expression levels were normalized as log2 values using GeneSpring software (Agilent Technologies, Santa Clara, CA, USA), and the obtained list contained genes that were up- or downregulated with greater than twofold changes in response to KSRP-KD. miRNA OneArray® and data analysis To identify which miRNAs are regulated by KSRP, a highthroughput and specific miRNA microarray (human miRNA OneArray® miRNA profiling chip) using Caki-1 cells with/without KSRP-KD was conducted by the Phalanx Biotech Group (Hsinchu, Taiwan).
RNA Amplification:
other:
Isolation:Article Title: Anti-Diabetic Effect of Lactobacillus Paracasei Isolated from Malaysian Water Kefir Grains.
Article Snippet: The prevalence of type 2 diabetes mellitus (T2DM) is alarming because it is always linked to the increase in chronic diseases, mortality, and socioeconomic burden.. Water kefir has a wide range of functional and probiotic characteristics attributed to the microorganisms present in the kefir grains.. The present study aims to evaluate the in vivo anti-diabetic potential of the isolated Lactobacillus paracasei from Malaysian water kefir grains (MWKG) which was reported to have excellent probiotic properties and high antioxidant activities as reported previously.
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