Silkworm strain that cocoons are reeled without using the boiling water may provide not only cost cut but also new material. It was clear that the water reeling strain which was bred in our laboratory was affected on the environment during the spining stage same as a general varieties, and it’s main factor was impossible to grope for fiber ends. We examined effect of humidity which affect the reelability on grouping end efficiency in the water reeling strains. N43 (from J 02, water reeling strain), N17 (from Daizo) and their Hybrid, N17×N43 were used in the experiment, 4 humidity conditions (low, midA, midB, high) were provided. The hybrid of N17×N43 was able to make cocoon at the experiment. On the other hands, 80% of N43 could not make cocoons and died without pupation at high humidity condition (99% RH). N17 which was another parent of hybrid was investigated in next raring season. Consequently, any larvae did not make a cocoon and some naked pupae appeared in N17. It may be suggested that hybrid larvae of N17×N43 made cocoons by heterosis at high humidity condition (99%RH).
This method is reported for the cryopreservation of ovaries and spermatozoa of the transgenic silkworm (Bombyx mori). At first, female moths were artificially inseminated with frozen-thawed semen of transgenic silkworm strains. The fertility varied from 0% to 92.0% depending on the strain. Addition of frozen-thawed semen from triploid moths, which are infertile but include intact apyrene sperm, improved fecundity of cryopreserved semen. The second experiment was ovary transplantation. Ovaries were removed from 5th instar larvae and cryopreserved in liquid nitrogen. The frozen-thawed ovaries were transplanted to 4th instar female larvae which had been surgically castrated. The emerged female moths were mated with nontreated male moths and deposited eggs. The eggs were genetically identical to the donor transgenic silkworm strains. Eggs were surgically removed from ovarian tubules of moths following transplantation of frozen-thawed ovaries and subjected to parthenogenetic activation and artificial hatching. All resulting moths were female and artificially inseminated with frozen-thawed semen. The used semen was collected from the same transgenic silkworm strains. Then the female moths yielded sound offspring of the transgenic silkworm strain at high rates. The third experiment was method of connecting the thread bodies of the donor with those of ovaries of the recipients. The results of connecting the thread bodies of ovaries were relatively high percentage. The emerged female moths artificially inseminated with frozen-thawed semen yielded well-conditioned offspring of the transgenic silkworm strains.
Recently, various kinds of transgenic silkworms, Bombyx mori, which spin innovative silk, such as fluorescent or extremely thin one, have been developed. Practical application of the value-added silkworms can be a major solution to revitalize sericultural farmers, who confront low cocoon price. The use of transgenic organism is regulated by the Cartagena law in Japan on the purpose to conserve biodiversity. Silkworm itself is a highly domesticated animal which is unable to feed or reproduce under natural conditions without human care. It is unlikely that the introduction of the genes encoding genetically engineered silk enables the silkworm survive independently to become a threat to biodiversity through competition with wild animals or feeding on natural plants. The wild silkworm, B. mandarina, inhabiting Japan can cross with B. mori artificially but no hybrid has been identified in the natural population of Japan. One reason of the absence of hybrids is that moths do not emerge in sericultural farms because pupae are killed in their cocoons for silk reeling. It is also supposed that the silkworms discarded outdoor cannot grow to mate with their wild relatives. In this study, we found that both larvae and moths of B. mori were preyed rapidly under natural conditions by various predators, such as birds, wasps, ants, and carrion beetles. We also developed methods for the risk assessment of rearing transgenic silkworm on biodiversity and applied them to a transgenic silkworm strain producing fusion protein of fibroin and green fluorescent protein (GFP) of jellyfish (Aequorea victoria). In the measurement of walking distance of individual fifth-instar larvae in 16hours, no statistical difference was observed between transgenic and non-transgenic silkworms. We also compared the position of eggs laid by individual female moths after mating and obtained a result that transgenic silkmoths laid eggs in smaller area than non-transgenic ones. The production of allelochemical substances was also tested by biological assay. On the soil mixed with freeze-dried powder of larval body or feces, there were no statistical differences between transgenic and non-transgenic silkworms for the rate of germination or weight of the seedling. The number of bacteria and fungi was also compared after the incubation of the soil mixed with the powdered specimens, resulting in no statistical differences between transgenic and non-transgenic silkworms in most of the tests. In combination with conventional statistics in silkworm breeding, such as larval days, cocoon weight, and egg numbers, researchers can assess the risk of their transgenic silkworm on biodiversity for the application of permission to rear them in sericultural farms