Mettl14

Manufactured by GenePharma

Sourced in China

METTLER14 is a laboratory equipment used for molecular biology and genomics research. It is a component of the methyltransferase complex that catalyzes the addition of methyl groups to RNA.

Automatically generated - may contain errors

Lab products found in correlation

Mettl3, by GenePharma (6 mentions) Lipofectamine 3000, by Thermo Fisher Scientific (3 mentions) Alkbh5, by GenePharma (2 mentions) Ythdf1, by GenePharma (2 mentions) Puromycin, by Solarbio (2 mentions) Igf2bp2, by GenePharma (2 mentions) Ythdc1, by GenePharma (1 mentions) Rpmi 1640 medium, by Solarbio (1 mentions) Fetal bovine serum (fbs), by Solarbio (1 mentions) Sgc 7901, by National Collection of Authenticated Cell Cultures (1 mentions) Hgc 27, by National Collection of Authenticated Cell Cultures (1 mentions) Ges 1, by National Collection of Authenticated Cell Cultures (1 mentions) Mkn 45, by National Collection of Authenticated Cell Cultures (1 mentions) Rnase r, by Yeasen (1 mentions) Sirna, by GenePharma (1 mentions) Pcdh ef1α mcs t2a puro, by System Biosciences (1 mentions) Plvx shrna2 bsd lentiviral vector, by Takara Bio (1 mentions) Lipofectamine 3000 reagent, by Thermo Fisher Scientific (1 mentions) Shrna nc, by GenePharma (1 mentions)

9 protocols using mettl14

Manipulation of Circular RNA Pathways

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Circular RNA NRIP1, ALKBH5, WTAP, YTHDF1, FTO, METTL3, METTL14, and NC siRNAs were obtained from GenePharma and their sequences are shown in Table S2. The inhibitors and mimics of miRNAs are shown in Table S3. Lipofectamine 3000 (Invitrogen) was used for transfections, following the manufacturer's instructions. A recombinant lentivirus containing sh‐circNRIP1 or sh‐ALKBH5 was established to stably knockdown circNRIP1 or ALKBH5. A nontargeting shRNA was used as the negative control. Circular RNA NRIP1 or ALKBH5 were overexpressed using recombinant lentiviruses containing complete coding sequences of these genes.
An empty lentivirus (vector) was used as the negative control. Lentiviral vectors were constructed by Obio Technologies. Infected cells were selected using puromycin. Quantitative real‐time PCR and immunoblotting were carried out to assess the transfection and infection efficiencies.

Ji X., Lv C., Huang J., Dong W., Sun W, & Zhang H. (2023). ALKBH5‐induced circular RNA NRIP1 promotes glycolysis in thyroid cancer cells by targeting PKM2. Cancer Science, 114(6), 2318-2334.

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Modulating IGF2BP3, SCD, and METTL14 in Cells

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We bought small interfering RNAs (siRNAs) targeting IGF2BP3, SCD, and methyltransferase-like 14 (METTL14) from GenePharma (Shanghai, China). SiRNA and negative control (NC) siRNA was transfected with Lipofectamine 3000 (Invitrogen, USA) ~48 h. qRT-PCR and western blotting were used to verify the transfection efficiency.
Next, the pHBLV-U6-MCS-EF1-Luc-T2A-Puro vector (HANBIO, Shanghai, China) inserted IGF2BP3-shRNA lentivirus, then the lentiviruses-infected CC cells grew in DMEM containing 5 µg/ml polybrene then. IGF2BP3-knockdown cells were transfected with the SCD-overexpression plasmid to rescue. Stable cell lines were produced by selection using 10 µg/ml puromycin (Solarbio, China). The aforesaid sequences appear in Table S1.

Han C., Hu C., Liu T., Sun Y., Hu F., He Y., Zhang J., Chen J., Ding J., Fan J., Zhang X., Wang J., Qiao X., Jiang D., Yang K, & Yang S. (2024). IGF2BP3 enhances lipid metabolism in cervical cancer by upregulating the expression of SCD. Cell Death & Disease, 15(2), 138.

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siRNA knockdown of key genes in RA-FLSs

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The small interfering RNAs (siRNAs) of IGF2BP2, GSTM5, METTL3 and METTL14 were obtained from GenePharma (Shanghai, China). The sequences are as follows: si-IGF2BP2-1 sense: 5′-GCGAAAGGAUGGUCAUCAUTT-3′; si-IGF2BP2-2 sense: 5′-GCUGUUAACCAACAAGCCATT-3′; si-IGF2BP2-3 sense: 5′-ACAGGACUGUCCGUGCUAUTT-3′; si-GSTM5-1 sense: 5′-GCUGGUCAGACUGUGCUAUTT-3′; si-GSTM5-2 sense: 5′-GGAUUCCUUGCCUAUGAUTT-3′; si-GSTM5-3 sense: 5’-GGGUUUGAAGAAGAUCUCUTT-3′; si-GSTM5-4 sense: 5′-GAAAGUCAGCUACAUGGAATT-3′; si-METTL3 sense: 5′-GCUGCACUUCAGACGAAUUTT-3′; si-METTL14 sense: 5′-GCAGCACCUCGAUCAUUUATT-3′. The siRNAs were transfected into RA-FLSs using a transfection reagent according to the manufacturer’s recommendations. At 6 h after transfection, the medium was replaced with a complete medium. The cells were collected for subsequent experiments at 48 h post-transfection.

Nan Y., Chen M., Wu W., Huang R., Sun W., Lu Q., Gu Z., Mao X., Xu H, & Wang Y. (2024). IGF2BP2 regulates the inflammation of fibroblast-like synoviocytes via GSTM5 in rheumatoid arthritis. Cell Death Discovery, 10, 215.

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Genetic Manipulation of Gastric Cancer Cells

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The normal gastric epithelial cell line GES-1 and GC cell lines (AGS, HGC27, MKN45, and SGC-7901) were purchased from the Cell Bank of the Chinese Academy of Science (Shanghai, China). All cell lines were incubated with RPMI 1640 medium (Solarbio, Beijing, China) supplemented with 10% FBS (Solarbio) in a humidified incubator at 37 °C containing 5% CO2.
FAM120A, WTAP, METTL3, METTL14, YTHDC1, SLCTA11, and PD-L1 knockdown and overexpression lentiviruses and their respective control vectors were generated by GenePharma (Shanghai, China). GC cells were inoculated into 6-well dishes and then infected with lentivirus at 60% confluence. Stable transfected cells were produced by treating cells with puromycin (4 μg/ml) for 2 weeks. The shRNA sequences are listed in Supplementary Table 2.

Niu L., Li Y., Huang G., Huang W., Fu J, & Feng L. (2024). FAM120A deficiency improves resistance to cisplatin in gastric cancer by promoting ferroptosis. Communications Biology, 7, 399.

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RNA and Protein Modulation Protocol

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RNase R (20 U/µl, Yeasen) was used to treat total RNA (20 µg) at 37 °C for 15 min. DNA transcription and replication inhibitor actinomycin D (Act D, HY-17,559), protein synthesis inhibitor cycloheximide (CHX, HY-12,320), and vimentin inhibitor Withaferin A (WFA, HY-N2065) were purchased from MedChemExpress (Shanghai, China). SiRNAs targeting ARIH1, vimentin, FTO, METTL3, and METTL14 were purchased from GenePharma (Jiangsu, China).

Zhang Y., Liu Z., Zhong Z., Ji Y., Guo H., Wang W, & Chen C. (2024). A tumor suppressor protein encoded by circKEAP1 inhibits osteosarcoma cell stemness and metastasis by promoting vimentin proteasome degradation and activating anti-tumor immunity. Journal of Experimental & Clinical Cancer Research : CR, 43, 52.

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Knockdown and Overexpression of TRIM9 and METTL14

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Short hairpin (sh)RNA targeting TRIM9 for knockdown (Sh‐TRIM9), shRNA targeting METTL14 for knockdown (METTL14‐KD) and the respective negative controls (NC) (Shanghai GenePharma). The shRNA sequence Sh‐TRIM9 and METTL14‐KD were constructed into the pLVX‐shRNA2‐BSD lentiviral vector (Takara Bio USA, Inc.). Overexpression TRIM9 (Oe‐TRIM9) and overexpression METTL14 (Oe‐TRIM14), lentiviruses (System Biosciences, LLC) were constructed into the lentiviral vector pCDH‐EF1α‐MCS‐T2A‐Puro (System Biosciences, LLC). Cells in each well were transfected with 100 nM miR‐29c‐3p mimic or 200 nM miR‐29c‐3p inhibitor.

Wu H., Jiao Y., Guo X., Wu Z, & Lv Q. (2024). METTL14/miR‐29c‐3p axis drives aerobic glycolysis to promote triple‐negative breast cancer progression though TRIM9‐mediated PKM2 ubiquitination. Journal of Cellular and Molecular Medicine, 28(3), e18112.

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RNA Interference Targeting Epigenetic Regulators

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METTL3, METTL14, IGF2BP1/2/3 and IFIT1 small interfering RNAs (siRNAs) were obtained from GenePharma (Shanghai, China). IFIT1 lentivirus shRNAs and pcDNA3.1_IFIT1 overexpressed lentivirus were also obtained from GenePharma (Shanghai, China). The control plasmid, METTL3, METTL14, IFIT1 and PD-L1 overexpressing plasmids were constructed in Generay Technologies (Shanghai, China). Cells were seeded in six-well plates overnight. For siRNA transfection, we made the transfection mixture with 5ul siRNA(50nM), 5ul Dharma FECT 1 transfection reagent and 390ul opti-MEM medium. For plasmid transfection, we added 1ug plasmid and 3ul viaFECT for one well. After six hours of incubation, the medium was replaced by normal culture media. Cells were harvested after 48h. For lentivirus shRNA transfection, cells were incubated with lentivirus at the MOI of 100. After 24 hours’ incubation, cells were treated with puromycin for two weeks. The sequences of siRNAs and shRNAs were listed in Supplementary Table 7.

Gao Y., Zou T., Xu P., Wang Y., Jiang Y., Chen Y.X., Chen H., Hong J, & Fang J.Y. (2022). Fusobacterium nucleatum stimulates cell proliferation and promotes PD-L1 expression via IFIT1-related signal in colorectal cancer. Neoplasia (New York, N.Y.), 35, 100850.

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Silencing HPV and IGF2BP2 in Cervical Cancer

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Small interfering RNAs (siRNAs) targeting HPV16 E6/E7, HPV18 E6/E7, IGF2BP2, and methyltransferase-like 14 (METTL14) were purchased from GenePharma (Shanghai, China). siRNA and negative control (NC) siRNA transfection was performed with Lipofectamine 3000 reagent (Invitrogen, USA) for 48 h. The transfection efficiency was verified by qRT-PCR and western blotting.
The HPV16 E6/E7-shRNA lentivirus, HPV18 E6/E7-shRNA lentivirus, and NC-shRNA were constructed by GenePharma (Shanghai, China). LV17-NC and LV17-Homo IGF2BP2 were cloned into the pcDNA3.1 (+) vector. The CC cells were infected with lentiviruses and cultured in DMEM supplemented with 5 µg/ml polybrene. To rescue the effect of E6/E7 knockdown on CC cells, HPV16/18 E6/E7-knockdown cells were infected with the IGF2BP2-overexpression lentivirus. Stable cell lines were generated by selection with 10 µg/ml puromycin (Solarbio, China). The expression of HPV16 E6/E7, HPV18 E6/E7, and IGF2BP2 was validated by qRT-PCR and western blot. The siRNA, shRNA and plasmid sequences are listed in Table S1.

Hu C., Liu T., Han C., Xuan Y., Jiang D., Sun Y., Zhang X., Zhang W., Xu Y., Liu Y., Pan J., Wang J., Fan J., Che Y., Huang Y., Zhang J., Ding J., Yang S, & Yang K. (2022). HPV E6/E7 promotes aerobic glycolysis in cervical cancer by regulating IGF2BP2 to stabilize m6A-MYC expression. International Journal of Biological Sciences, 18(2), 507-521.

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Modulating m6A Regulators in Cells

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CircNRIP1, ALKBH5, WTAP, YTHDF1, FTO, METTL3, METTL14 and NC siRNAs were obtained from GenePharma (Suzhou, China) and their sequences are shown in Additional le 1 (Table S2).
Lipofectamine 3000 (Invitrogen, Waltham, MA, USA) was used to perform transfections, according to the manufacturer's instructions. A recombinant lentivirus containing sh-circNRIP1 or sh-ALKBH5 was established to stably knock out circNRIP1 or ALKBH5. A non-targeting shRNA (sh-NC) was used as the negative control. circNRIP1 or ALKBH5 were overexpressed using recombinant lentiviruses containing the complete coding sequences of these genes.
An empty lentivirus (vector) was used as the negative control. The lentiviral vectors were constructed by Obio Technologies (Shanghai, China). Infected cells were selected using puromycin. Then, qRT-PCR and immunoblotting were performed to assess the transfection and infection e ciencies.

Ji X., Wang W., Lv C., Huang J., & Zhang H. (2022). ALKBH5-induced circRNA NRIP1 promotes glycolysis in thyroid cancer cells by targeting the miR-541-5p/PKM2 and miR-3064-5p/PKM2 axes.

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