4.8. Transcriptome Measurement

After extracting total RNA, the quality and quantity of RNA were detected by Nanodrop and Agilent. Qualified RNA was processed for library construction. The raw data were generated using the Illumina NovaSeq 6000 platform as PE150. After the raw reads were filtered, 5.74 GB of clean data and above the 95.24% of Q30 bases per sample were obtained in the first experiment, while 5.81 GB of clean data and above the 91.98% of Q30 bases in each sample were acquired in the second experiment of leaf development. Fastp software was used to control the quality of clean data [70]. The genomic index of P. alba was built and the cleaned reads were mapped to the genome via hisat2 [71]. Then, Samtools software compressed and sorted the mapped data [72]. The read counts and TPM of each gene were calculated by R script. The R packages, including DESeq2, clusterProfiler and ggplot2, were used for differential expression genes analysis and plot drawing. In the first stage experiment, the fold change > 1.5 and FDR < 0.05 were both regarded as the threshold for filtering DEGs.