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Eukaryotic mRNA Sequencing with Reference Genome

FAQ

  •  A brief introduction of the construction of eukaryotic transcriptome library


    1) mRNA was separated by magnetic beads for polyA structure of mRNA

    2) mRNA fragmentation

    3) The base T is replaced by U during the second strand of cDNA synthetization to achieve the purpose of strand specific library.

    4) 5 'end repair, 3' end add “A”

    5) Add adapters

    6) Collect fragments with specific length, generally 300bp

    7) PCR

    8) Sequencing


  • How to select appropriate genes for qRT-PCR verification?


     Firstly, according to the enrichment results of GO/KEGG, the genes concerned shall be selected. Then, according to the results of differential expression analysis, the genes with higher fold change value, smaller P value and higher FPKM value shall be verified by qRT-PCR.