Maize B73 seeds were germinated in hydroponic culture under sterile conditions. After germination, we moved seedlings of maize B73 to a sand-based pot culture to inoculate them with arbuscular mycorrhizal fungus. The control group was not inoculated with arbuscular mycorrhizal fungi. Culture medium was mixed with vermiculite:perlite at a 4:1 ratio and sterilised by 40 min high-pressure steam at 121℃. The AMF species was Glomus intraradices (Gi, provided by Sun Yat-Sen University). We grew maize at 28°C with 16 hours of light and 8 hours of darkness in a greenhouse. After 60 days post-treatment, the plants were collected. For RNA isolation, samples were stored at 80°C.
TransZol Up Plus RNA Kit was used to extract total RNA (TransGen Biotech, Beijing, China). cDNAs were obtained using reverse transcriptase (Vazyme, Nanjing, China). ZmTubulin (Zm00001d009780) and ZmGAPDH (Zm00001d049641) were used as the endogenous reference genes to standardize the relative expression levels of ZmGRF genes. Our previous published work provided extensive knowledge regarding qRT-PCR reactions (Wang et al., 2022 (link)). The 10-(ΔCt/3)method was used to determine the relative gene expression levels (Li et al., 2005 (link); Liao et al., 2014 (link)). The qPCR assays were performed with three biological replicates.
TransZol Up Plus RNA Kit was used to extract total RNA (TransGen Biotech, Beijing, China). cDNAs were obtained using reverse transcriptase (Vazyme, Nanjing, China). ZmTubulin (Zm00001d009780) and ZmGAPDH (Zm00001d049641) were used as the endogenous reference genes to standardize the relative expression levels of ZmGRF genes. Our previous published work provided extensive knowledge regarding qRT-PCR reactions (Wang et al., 2022 (link)). The 10-(ΔCt/3)method was used to determine the relative gene expression levels (Li et al., 2005 (link); Liao et al., 2014 (link)). The qPCR assays were performed with three biological replicates.
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