The internal reference gene developed based on Miscanthus transcriptome sequence and its application
A technology of transcriptome sequence and internal reference gene, which is applied to the internal reference gene developed based on Miscanthus transcriptome sequence and its application field, can solve the problems that restrict the process of gene screening and verification research, and achieve abundant available quantity, good specificity and stability sex high effect
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Embodiment 1
[0059] This example is the comparative application of the three internal reference genes developed based on the Miscanthus transcriptome sequence described in the present invention and nine traditional internal reference genes in the gene analysis of Miscanthus under drought stress. In order to compare the effects of developing new internal reference genes, the following seven The commonly used traditional reference genes GAPDH, 18S rRNA, ACTIN, PP2A, eIF4a, SAMS, SAMDC and the reference genes Sb09g019750 and Sb02g041180 on two closely related species sorghum were used as controls. The sequence information of common internal reference genes was obtained from the NCBI database, and primers were designed online using primer3.0 after homologous gene sequence alignment. The primer-related information is shown in Table 4.
[0060] Specifically include the following steps:
[0061] 1), material selection: select Miscanthus seeds, and sterilize the selected seeds with 75% alcohol and...
Embodiment 2
[0082] In this example, in order to verify the effect of the application of the internal reference gene, two genes, Cu / Zn SOD and CAT, were selected as the target genes. SOD and CAT are common antioxidant enzymes in plants, which can help plants remove reactive oxygen species in the body and protect cells from immune cells. Oxidative damage usually increases its expression after stress. Taking Unigene33312 and common reference gene ACTIN as examples to analyze the expression characteristics of awn SOD and CAT genes after drought stress, the specific steps are as follows:
[0083] 1), material selection: Select Miscanthus (two varieties: 0819 and 1302) seeds, and sterilize the selected seeds with 75% alcohol and 1% sodium hypochlorite. The experimental seeds were germinated in a petri dish and then transplanted into plastic pots (9cm in diameter) for sand culture, and 1 times of Hoagland nutrient solution was poured into the incubator for cultivation. Circulation, relative hum...
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