Hi Abda, I have not seen any baculavirus vector for larvae expression. However, if you have a protocol for transfection by using larvae, I imagine any insect virus system would work, such as virus generated by MultiBac etc. It is nice to see you are from Huazhong Agricultural Univ.
Hi,
I used insects cells transfected by bacculovirus ( with gene of Glycoprotein, Nucleoprotein or the both, and native cells ( against rabies, I used this cells on mice I.P, goat and dogs I.M . our results are very interessant ( it was be communicated on 13 eme Journées internationales de Biotechnologie (JIB 2014) : 20- 24 december 2014).
I worked for a larvae-based baculovirus expression vector company for 4 years. It is in some ways more challenging than cell culture but has some advantages in costs and sterility requirements.
The same baculovirus systems that are used for insect cell culture are used for insect larvae. BacPAK, Bac-to-Bac, AB Vector and Flashbac will all infect insect larvae. One needs to use the correct species of insect and only infect the larval stage. Cabbage looper (Trichoplusia ni) are good for producing proteins from the most commonly used baculovirus expression vector systems.
In all cases, larvae can be inoculated by injection budded virus from the insect cell culture media. A Hamilton syringe is good for doing injections. Third instar larvae are a good size to inject if it is T.ni.
Larvae can also be fed baculovirus-infected insect cells which have been homogenized without detergent, suspended in water and spread across the insect diet. The infected cells contain pre-occluded virions that are orally infectious to larvae. The media is not orally infectious to larvae.
If you're trying to do things without access to insect cell culture, liposome, lipofectamine or PEI based transfection reagents will work fine to transfect insect larvae. The best baculovirus vector option in this case is the Bac-to-Bac bacmid system. Purified bacmid DNA can be transfected into insect larvae by injection. Amounts to inject can be determined empirically. No need to find a protocol on this.
Once the larvae have died from the infection, they can be harvested and lyophilized to become inoculums of thousands of larvae. Lyophilization of larvae is an excellent way to store inoculum as it can be ground into a powder, mixed with water and applied to diet.
Getting some experience rearing insect larvae is recommended before trying any of this. It is also helpful to make a DS-red or GFP recombinant baculovirus to optimize the infection and initial transfection conditions.
One final tip. Add 1mM DTT or BME when homogenizing larvae or else everything will turn black.
Best of luck.
- Badges
- Science topic
- Similar topics
- Biological Science
- Zoology
- Invertebrate Zoology
- Arthropods
- Insects
- Insect