Oocyte RNA Extraction and qPCR

As described in our previous study (14 (link)), total RNA was extracted from single oocytes using the SMARTer Ultra Low RNA kit for Illumina sequencing. Synthetic cDNA was used for quantitative real-time PCR (qPCR) with an ABI 7500 PCR machine. Gene expression was normalized to the expression of the housekeeping gene GAPDH. Primers were synthesized by Sangon Biotech (Shanghai) Co., Ltd. For mRNA expression identification, 16 IVM and 16 IVO human oocytes were used to extract RNA from each oocyte, respectively.

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Zhao H., Li T., Zhao Y., Tan T., Liu C., Liu Y., Chang L., Huang N., Li C., Fan Y., Yu Y., Li R, & Qiao J. (2018). Single-Cell Transcriptomics of Human Oocytes: Environment-Driven Metabolic Competition and Compensatory Mechanisms During Oocyte Maturation. Antioxidants & Redox Signaling, 30(4), 542-559.

Publication 2018

SMARTer Ultra Low RNA kit

Cdna Gapdh Gene expression Housekeeping gene Human Mrna Oocyte Primers Quantitative real time pcr

Corresponding Organization : Guangzhou Medical University

Other organizations : Kunming University of Science and Technology

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Variable analysis

independent variables

Culture method: IVM (in vitro maturation) vs. IVO (in vivo maturation)

dependent variables

Gene expression of target genes

control variables

Housekeeping gene GAPDH for normalization of gene expression
Quantitative real-time PCR (qPCR) using an ABI 7500 PCR machine
RNA extraction using the SMARTer Ultra Low RNA kit for Illumina sequencing
Number of oocytes used: 16 IVM and 16 IVO human oocytes

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

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