关键词: 麦长管蚜, miRNA; U6, 内参基因, 翅型分化, 表达谱

Abstract: 【Aim】 This study aims to clarify the expression profiles of nine microRNAs (miRNAs) in the grain aphid, Sitobion avenae at different developmental stages between winged and wingless morphs, and to define the key stage of the aphid wing dimorphism. 【Methods】The full-length cDNA of the reference gene U6 and nine miRNAs of S. avenae were cloned by using RT-PCR. The expression levels of nine miRNAs were detected in winged and wingless S. avenae at different developmental stages (pseudo embryo, 1st-4th instar nymph and adult) by using qRT-PCR. 【Results】The full-length cDNA of U6 of S. avenae is 92 bp and shows over 80% nucleotide sequence identity with U6 from other reported insects. qRT-PCR results revealed that most miRNAs had low relative expression levels in the pseudo embryo and nymphal stages, and high relative expression levels at the adult stage. The relative expression levels of six miRNAs (miR-277, miR-9a, miR-7, Let-7, miR-1 and miR-315) in winged adults were higher than those in wingless adults. MiR-277 and miR-9a had higher relative expression levels in winged adults. Except miR-315, the relative expression levels of five miRNAs including miR-277, miR-9a, miR-7, Let-7 and miR-1 in S. avenae at the adult stage showed significant difference between the winged and wingless morphs. Moreover, there were significant differences in the relative expression levels of miR-9a in the pseudo embryo and miR-315 in the 3rd instar nymphal stage between the winged and wingless morphs. The other three miRNAs (miR-8, PC-3p-2743_844 and PC-5p-113190_15) showed higher relative expression levels in wingless adults than in winged adults. The relative expression levels of miR-8 and PC-3p-2743_844 in winged and wingless adults were significantly higher than those in the other developmental stages, and that of PC-5p-113190_15 in the 3rd nymphs of wingless aphid was significantly higher than those in the other developmental stages. The relative expression levels of miR-8 in the adult stage and PC-3p-2743_844 in the pseudo embryo and adult stages showed significant difference between the winged and wingless morphs, and that of PC-5p-113190_15 in the 3rd nymphal stage showed extremely significant difference between winged and wingless morphs. 【Conclusion】The reference gene U6 of S. avenae is highly conserved among insects. The expression profiling results suggest that nine miRNAs may regulate the wing-morph differentiation of S. avenae during the pseudo embryo and adult stages.

Key words: Sitobion avenae, miRNA, U6, reference gene, wing dimorphism, expression profile