Method for relieving diapause of silkworm diapause cysts through very early coronas

Abstract

The invention relates to a method for preparing a silkworm diapuse variety of transgenic silkworms by means of very early coronas. The method comprises the steps that diapuse silkworm eggs obtained 2-4 hours after being laid are subjected to corona treatment for 30-120 seconds in an electric field with the voltage strength of 8-12 kV, then the silkworm eggs obtained after corona treatment are sterilized, microinjected, sterilized again and incubated in sequence, the G0 generation of silkworm moths are obtained, the obtained G0 generation of silkworm moths are subjected to selfing or backcrossing, and the G1 generation of silkworm eggs are obtained; transgenic individuals are screened, and the transgenic variety of the silkworm diapuse variety is obtained through raising and passage of the transgenic individuals. By means of the method, the bottleneck of the silkworm diapuse variety transgenic technology is effectively broken through, and the method has an important significance on quickly achieving the silkworm transgenic industrialization.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for releasing the diapause of silkworm eggs by very early corona, and also relates to silkworm eggs released by very early corona and a method for preparing transgenic silkworms by using the released silkworm eggs. Background technique [0002] Transgenic technology is an important technology to realize gene function verification and genetic improvement in modern biological research. In the post-genome era, transgenic technology has matured and played a huge role in functional genome research, directed genetic breeding, establishment of human disease models, and bioreactor development. [0003] The silkworm is one of the most successful and longest economic insects used by humans in the world. The domestication and breeding of the silkworm have a history of about 5,000 years. Bombyx mori is also a model of Lepidoptera insects. The completion of silkworm genome f...

AI Technical Summary

Problems solved by technology

In summary, the use of polymorphic silkworm varieties as transgenic breeding materials has two obvious defects: ① The economic traits of polymorphic silkworm varieties are poor, and the additional traits created by transgenes cannot be directly popularized and applied in terms of yield and quality According to the standards, the transgenic strains with polymorphic silkworm eggs as raw materials can only be used as raw materials for breeding, and as the original carrier of a certain relevant excellent trait, they can be used in the long and arduous traditional breeding

②Pluripotent silkworm varieties do not have a diapause period, and the maintenance of their subculture can only rely on continuous breeding and reproduction, which wastes a lot of manpower and material resources, and increases the risk of resource loss of transgenic silkworm varieties obtained

Although the above three patents have respectively provided corresponding transgenic methods for diapause species, which can also realize the transgenesis of most diapause species, the methods provided by them have their own limitations and shortcomings: the early pickling transgenic method of silkworm diapause species (CN101195834A) Due to the need for microinjection immediately after pickling, two consecutive stimulations to silkworm eggs resulted in extremely low hatching rates after injection, and the effect of early pickling varies greatly among varieties, so it is mainly suitable for acid-resistant stimulation. Japanese varieties

Although the low-temperature acceleration greening transgenic method of silkworm diapause species (CN101195833A) has the same hatching rate after injection as the polymorphic silkworm eggs, the effect of low-temperature acceleration greening is very different among varieties, and its main adaptability changes are affected by temperature. The Chinese varieties that are more sensitive to the impact are less effective in relieving diapause for monomorphic and Japanese dimorphic silkworm varieties

In addition, due to the long period of low-temperature acceleration of the parent silkworm eggs and the poor physical fitness of the non-diapause offspring silkworms obtained after low-temperature acceleration, it seriously affects the survival rate of silkworms after injection hatching.

While the improved method for transgenic silkworm diapause at low temperature (CN101503704A) has improved the diapause effect of Japanese dimorphic silkworm varieties, it still has the same transgenic method as the low-temperature acceleration of silkworm diapause (application number: CN101195833A). Shortcomings

These shortcomings greatly limit the large-scale application of silkworm transgenic technology in practical research such as functional gene research and bioreactor development.

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