| Major royal jelly protein, one of the most important protein components in royal jelly, play an very important role in bee biology and beekeeping.There are 10 members in MRJPs. Among them the MRJP1-5 in the royal jelly content is very high, while the content of the remaining family members are rarely even undetectable. In order to explore the gene expression and biological function of mrjp6 and mrjp8, the expression of mrjp6 in different development stages and different tissues of three castes in honeybee were determined. At the same time, The expression of mrjp8 was inhibited by using RNA interface technology. The specific results are as follows:Firstly, to clarify the temporal and spatial expression of gene mrjp6 in honeybee(Apis mellifera L.) for providing a basis for further biological function research. In this study, real-time PCR was employed to determine the mrjp6 expression in honeybee(Apis mellifera L.) with different caste, developmental stages and tissues. The result showed that the mrjp6 was low expressed in larva and pupae stages, which was almost identical to the control; Its expression increased significantly in adult honeybee, which was 25～28 times of control (p<0.05). In adult bee, mrjp6 was low expressed in drone bee, which was 25 times of control, but significantly higher expressed in female honeybee including worker and queen, which was up to 28 times of control (p<0.05). These showed that the expression of mrjp6 was related to gender. However, the same expression level of mrjp6 between the adult queen and worker bee revealed that its expression was independent of caste. Further research showed that mrjp6 were stably high expressed in nurse and forager bee, and it was especially high expressed in head part by determining the expression of different tissues. As a member of MRJPs family, mrjp6 specifically high expressed in head of adult female bee (queen and worker), which was different with the typical express mode of the main MRJPs members, they were high expressed in the hypopharyngeal gland of nursing worker bee around 9 days old, but low expressed in other adult stages. Such result illustrated that the mrjp6 might has different biological function from the main MRJPs. We deduced that the mrjp6 may participate in these functions related to honeybee nervous system and the complicated behavior developmental of adult female honeybee.Secondly, to explore the biological function of mrjp8 in Apis mellifera L., dsRNA of mrjp8 were synthesised by two ways, the kit synthesize and the engineering bacteria synthesize, and then the synthesised dsRNA were used to interference the objected in worker and queen by either feeding or micro-injection, and the expression of mrjp8 was then quantifed after interference. The results show that the motrality level was 75% when feeding the worker and queen with 1 μg dsRNA. However, when the feeding amount of dsRNA decreased to 50 ng, the motrality level was not significant different among the three experimental groups. And the expression of mrjp8 was not affected when feeding the worker or queen with either 1 μg or 50 ng amount of mrjp8 dsRNA. The result of infection new born worker 1 μg mrjp8 dsRNA showed no difference what ever in gene expression level or protein level. Furthermore, an engineering bacteria containing the expressed dsRNA of mrjp8 were feeding to the honeybee for inhibited objected gene expression, and the result showed that the more bacteria fed, the higher motrality happened.But the expressing of mrjp8 was still not significant in dispite of a downward trend exhibited when fed at 4×109 concentration.In conclusion, The paper used qRT-PCR to quantitative the expression of mrjp6 in Apis mellifera. We found that the mrjp6 was significant differently expressed between the female (worker and queen) and male honeybee. It was highe experssed in female adult bees, especifically high expressed in worker bee’s head. Such result revealed that the mrjp6 might related with bee nervous system related fuction. While in mrjp8 dsRNA interference in Apis mellifera of, the result showed the expression of mrjp8 was not decreasing after feeding or injiecting dsRNA. The results implicated that mrjp8 may have an important role in honeybee development, and its function could not be replaced. The honeybee therefore has a corresponding mechanism to compensate for its expression, which might be the reason why it was difficult to be successful interfered. |
