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Sample GSM5114404

Query DataSets for GSM5114404

Status

Public on Mar 02, 2021

Title

F50 C2

Sample type

SRA

Source name

F50 silkworm

Organism

Bombyx mori

Characteristics

strain: F50
tag: day-6 fifth instar larvae
treatment: Control
tissue: prothoracic glands

Growth protocol

The silkworm strain F50 was provided by The Sericulture Research Institute, Chinese Academy of Agricultural Sciences (Zhenjiang, China). Then they were reared under standard conditions until spawning (25˚C and 70% humidity). Prothoracic glands (PGs) were carefully removed from the day-6 fifth instar larvae were of the infected and control groups after BmNPV infection 48 h. All samples were immediately stored in liquid nitrogen.

Extracted molecule

polyA RNA

Extraction protocol

Total RNA was extracted from PGs using TRIzol reagent (Invitrogen, New York, NY, USA) in accordance with the manufacturer’s instructions. The mRNA was purified by Oligo(dT) magnetic beads (Invitrogen, USA), then used as templates for cDNA synthesis.
Poly (A)-tailed RNA prepared using magnetic oligo (dT) beads was broken into short fragments using a fragment buffer and was then reverse-transcribed to synthesize the first-strand cDNA with a random primer, and then DNA polymerase I was mixed with RNase H, dNTP, and the buffer solution to synthesize the complementary strand. The libraries were constructed using the Illumina methods and protocols, following the manufacturer’s instructions. The insert size and concentration of the cDNA library were both checked and quantified by an Agilent Bioanalyzer 2100 (Agilent Technologies, Inc., Santa Clara, CA) and Qubit® RNA Assay Kit (Life Technologies, CA, USA), respectively. RNA-seq was carried out with an Illumina HiSeq 2500 instrument (Illumina, San Diego, CA, USA). In order to ensure the quality of information analysis, the raw reads were cleaned by removing the adapter sequences, empty reads, unknown nucleotides (ratio ≥10%), and low-quality reads with a basic mass value of Q ≤ 20 which accounted for more than 50% of the whole read length.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2500

Data processing

Quality analysis was conducted for the original data obtained using the Illumina HiSeqTM2500 system
Clean reads were screened by filtering out low-quality reads
A second quality analysis for alignment, analysis of the distribution and coverage of the clean reads on the reference sequence was conducted
The sequences were aligned to the silkworm genome database SilkDB.
FPKM of each gene were calculated based on the length of the gene and read count mapped to a given gene. The clean reads were mapped to the B. mori genome. Genes with a FPKM ≥1.0 were identified as expressions. A ratio (log2 fold change) between the infected and control groups of ≥1.5 was identified as the determinant of differentially expressed genes (DEGs).
Genome_build: SilkDB (http://silkworm.swu.edu.cn/silkdb/)
Supplementary_files_format_and_content: All files are named with sample name followed by content name using files format of excel. The gene and its log2 Ratio of TPM among all time points would be selected if it is a DEG in any comparison. The expression patterns were clustered using hierarchical cluster analysis. The Pearson's correlation of log2 Ratio among the genes is being used as the distance for clustering.

Submission date

Feb 27, 2021

Last update date

Mar 02, 2021

Contact name

Pingzhen Xu

E-mail(s)

xpz198249@163.com

Phone

15952901490

Organization name

Jiangsu University of Science and Technology