Alkbh5

Manufactured by GenePharma

Sourced in China

ALKBH5 is a laboratory equipment product that functions as a demethylase enzyme. It is involved in the regulation of N6-methyladenosine (m6A) modification in RNA molecules. ALKBH5 plays a role in the reversible methylation of RNA, which is an important post-transcriptional regulatory mechanism.

Automatically generated - may contain errors

Lab products found in correlation

Lipofectamine 3000, by Thermo Fisher Scientific (3 mentions) Mettl3, by GenePharma (2 mentions) Ythdf1, by GenePharma (2 mentions) Mettl14, by GenePharma (2 mentions) Scrambled sirna, by Thermo Fisher Scientific (1 mentions) Prrc2a sirna, by Thermo Fisher Scientific (1 mentions) Alkbh5 sirna, by Thermo Fisher Scientific (1 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions) Lipofectamine rnaimax, by Thermo Fisher Scientific (1 mentions) Lentiviral particles, by GenePharma (1 mentions)

5 protocols using alkbh5

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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Investigating Genes Regulating RNA Modification

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Mouse Prrc2a, Fto, and Alkbh5 siRNAs were designed and synthesized by Genepharma Corporation (Suzhou, China). The following siRNA were synthesized and used in the study: Prrc2a siRNA 1#: 5′-CAUGAAGAGGUUGACUAUA-3′, Prrc2a siRNA 2#: 5′-GCUUGUAUAUAGAUUAUAA-3′, FTO siRNA: 5′-GCAGCUGAAAUACCCUAAA-3′, Alkbh5 siRNA: 5′-ACAAGUACUUCUUCGGCGA-3′, Scrambled siRNA (siNC): 5′-UUCUCCGAACGUGUCACGU-3′. Transfections were performed with Lipofectamine RNAiMAX (Invitrogen) for siRNA, and Lipofectamine 2000 (Invitrogen) for plasmid following the manufacturer’s instructions.

Wu R., Li A., Sun B., Sun J.G., Zhang J., Zhang T., Chen Y., Xiao Y., Gao Y., Zhang Q., Ma J., Yang X., Liao Y., Lai W.Y., Qi X., Wang S., Shu Y., Wang H.L., Wang F., Yang Y.G, & Yuan Z. (2018). A novel m6A reader Prrc2a controls oligodendroglial specification and myelination. Cell Research, 29(1), 23-41.

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ALKBH5 Knockdown and Overexpression

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Lentivirus expressing scramble or ALKBH5 shRNAs was purchased from GenePharma Company. In the case of knockdown experiments, cells were infected these lentiviral particles and selected with 3 μg/ml puromycin. In the case of overexpression experiments, cells were infected with lentiviral particles expressing empty vector control or ALKBH5 (GenePharma Company) and selected with 3 μg/ml puromycin.

Chen S., Zhou L, & Wang Y. (2020). ALKBH5-mediated m6A demethylation of lncRNA PVT1 plays an oncogenic role in osteosarcoma. Cancer Cell International, 20, 34.

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Gene Expression Manipulation Protocols

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For gene overexpression, cells were transfected with empty vector or overexpressing vector of KCNK15-AS1, ALKBH5, KCNK15, or REST (GenePharma Company). For gene knockdown, sh/Ctrl, ALKBH5 shRNAs (sh/ALKBH5#1/2/3), PTEN shRNAs (sh/PTEN#1/2), and MDM2 shRNA (sh/MDM2) were acquired from GenePharma (Shanghai, China). All transfection was implemented via Lipofectamine 3000 (Invitrogen, USA) and transfection efficiency was verified using RT-qPCR.

He Y., Yue H., Cheng Y., Ding Z., Xu Z., Lv C., Wang Z., Wang J., Yin C., Hao H, & Chen C. (2021). ALKBH5-mediated m6A demethylation of KCNK15-AS1 inhibits pancreatic cancer progression via regulating KCNK15 and PTEN/AKT signaling. Cell Death & Disease, 12(12), 1121.

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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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