Research On Expression Of Circadian Rhythm Gene Period And Gene Timless In The Diapause-induce Process Of Silkworm, Bombyx Mori

Circadian rhythm is the most common behavior of organism in nature. Temperature and photoperiod are the two important zeitgebers that affect the expression of circadian gene in the nature.Bombyx mori is a model organism of lepioloptera, which has reached industrial production scale due to its economic importance.Diapause is a basic physiological phenomenon in B.mori and other insects.It is recognized as an important trait that affects the sericulture production and pest control.B.mori is a kind of insect with diapause during egg stage.Diapause inducements are due to temperature and photoperiod in the incubation process of parental eggs.It is of great theoretical value and practical significance to study the circadian molecular mechanism in the process of silkworm diapause,which can help to further illustrate the molecular mechanism of dispause in B.mori,and elevate our knowledge in pest prevention and control program.The latest homologous gene sequences of Gene period(per) and Gene timless(tim) of B.mori were from Dazao stain,and the ruction of the two genes was unknown at present.We investigated the expression profiles of the two genes throughout the generations(egg-larva-pupa-moth-baby egg) under photoperiod and temperature incubation conditions of parental egg using C108 strain as material.We investigated the circadian rhythm genes mechanism of action from the dispause regulation point.The offspring eggs are completely dispause under the 25℃and constant illumination incubation condition(25LL) of parental egg,and they are completely non-dispause under 15DD(15℃,constant dark),and mixture of dispause and non-dispause in 20LD(20℃,12h illumination and 12h dark).The per gene expression of eggs in 25LL is significantly higher than 15DD;it is also higher than 20LD.This phenomenon is even more obvious in the later incubation stage(7 days later) that the embryonic nervous system fully grown.The expression level of tim gene in 25LL is markedless higher than 15DD and 20LD in the early incubation stage(7 days before), but it is obviously higher than 20LD and 15DD conditions in the late.To further invesgate the per and tim expression in different moment every day of the sensitive period(7d,8d,9d) during the process of embryogeny,the expression of tim gene was very low and the per and tim expression did not have obvious rhythmicity under 15DD condition.But the expression of two genes was high and had obvious rhythmicity under 25LL and 20LD.In a word,after embryo nervous system has grown, the B.mori can strengthen the ability to perceive the illumination and temperature outside.Moreover,the two genes have different reflection to illumination and temperature.The studies of different tissues from Larva,Pupa to Moth had shown that photoperiod had no significant effects to the expression of per gene and tim gene in B. mori(1-3L for the whole silkworm) under different temperatures of 15℃or 25℃.At incubation temperature of 15℃,20℃and 25℃,expression of the two gene also had no significant differences in the larva,pupa and adult head,gonad and fat body under constant illumination or darkness.In temperature and illumination combinations,the expression of per and tim gene did not present remarkable rhythmicity.The above results indicated that photoperiod and illumination during incubation process had little effects on the expression of gene per and gene tim in the period from larvae to moth of B.mori.The offspring eggs of B.mori are all diapause due to the incubation condition of parental eggs under 25℃and constant illumination(25LL),after treated by hydrochloric acid when they were layed about 20h,per and tim always keep high transcription activity during the activation process of embryo.The expression of per and tim had no obvious differences in offspring eggs between different incubation temperatures 25℃and 15℃in constant illumination.The expression level of the two gene also had no significant differences in offspring eggs under constant light or darkness with low temperature 15℃.It suggested that temperature and photoperiod in egg incubation process may have no effects on the expression of gene per and tim of offspring eggs.Diapause induced zeitgebers(temperature and photoperiod) affect the expression of circadian genes per and tim in parental eggs directly during their incubation period but have no significant effect on the expression of the two genes in polypide and offspring egg.Take dispause-induce as phenotype,the zeitgebers(temperature and photoperiod) and the diapause induction condition as influencing factor,and the expression of period and timeless as result,we presume that circadian genes per and tim play an important role of upper location in the gene network of diapause inducement.