Mir 30b

TaqMan microRNA reverse transcription kit (Applied Biosystems) and high capacity RNA to cDNA kit were used as per the manufacturers' instructions to generate complementary DNA for miRNA and gene expression assays, respectively. RT-PCR was performed using TaqMan Universal PCR Master Mix (Applied Biosystems) for miRNA RT-PCR and Gene Expression Master Mix (Applied Biosystems) for mRNA quantification. RT-PCR assays were run on a Bio-Rad CFX96 Touch real time PCR detection system (Bio-Rad). The assays were performed in triplicate using either U6 (miRNA expression) or RPS9 (mRNA expression) as an internal control. All TaqMan assays were purchased from Applied Biosystems and include: miR-30a (assay 000417), miR-30b (assay 000602), miR-30c (assay 000419), miR-30d (assay 000420), miR-30e (assay 002223), miR-101a (assay 002253), miR-101b (assay 002531), miR-320a (assay 002277), miR-145 (assay 000467), U6 (assay 001973), Sox9 (assay Mm00448840_m1), Rps9 (assay Mm00850060_s1), SOX9 (assay Hs01001343_g1), HES1 (assay Hs00172878_m1), sucrose isomaltase (assay Hs00356112_m1), and RPS9 (assay Hs02339424_g1).

A TaqMan miRNA assay was used to detect the contents of miRNAs. The single-stranded cDNA was synthesized using the TaqMan miRNA reverse transcription kit (Applied Biosystems; Foster City, CA, USA) and then amplified by using the TaqMan universal PCR master mix (Applied Biosystems; Foster City, CA, USA) together with the following miRNA-specific TaqMan MGB probes: Let-7b, miR-125a, miR-181b, miR-16, miR-30b, and miR-101 (Applied Biosystems; Foster City, CA, USA).