| Silkworm ingests a large number of mulberry leaves to obtain rich nutrients to maintain subsequent growth and reproduction.Selenium has obvious nutritional value and biological regulation on animals.There are few reports on the regulation mechanism of selenium on the growth,development and reproduction of silkworm.In this thesis,the effects of different concentrations of selenium on the growth,development and reproduction of silkworm were investigated,and the enrichment capacity and distribution of selenium in silkworm tissues were determined.The changes of selenoprotein-related gene expression in silkworm were investigated after selenium supplementation.The physiological regulation of GPX gene on silkworm and the interaction between selenium supplementation and GPX gene interference were studied by RNA interference technique.The results provided a basis for the use of selenium to regulate the growth and development of silkworm.1.The effects of low concentration and high concentration feeding on the growth and development of silkworm were different.When selenium concentration is 50 μM,it can promote the growth of silkworm larvae.The average body weight during L5D1-L5D6 of the 50 μM group were 0.59±0.01 g,0.84±0.01 g,1.13±0.03 g,1.58±0.05 g,1.82±0.03 g and 2.16±0.04 g.In the 50 μM group,cocoon quality of silkworm was significantly improved.When the selenium concentration was 200 μM,the physiological condition of silkworm was negatively affected.The average body weight during L5D1-L5D6 of the 200μM group were 0.46±0.02 g,0.64±0.05 g,0.83±0.05 g,1.21±0.03 g,1.48±0.04 g and1.77±0.06 g at the same stage.When Se concentration was 200 μM,cocoon quality of silkworm decreased significantly.When selenium concentration was 200 μM,the reproductive capacity of silkworm was inhibited.In 200 μM group the larval stage of silkworm was prolonged by 36±7.27 h.The larval period of 50 μM group was prolonged by 15±3.42 h.2.Silkworm has the ability to enrich selenium and the selenium content is different in different tissues.When silkworm is exposed to the same selenium concentration,different organs of silkworm have different selenium enrichment abilities.The order was testis >head ≈ Malpighian tube > epidermis > fat body ≈ midgut > silk gland.The Se content in testis of control group were 2.169±0.248,50 μM group were 11.818±0.216 and 200 μM group 44.351±0.317 nmol/g FW.The selenium contents of head in control group,50 μM group and 200 μM group were 0.103±0.121,6.49±0.324 and 23.633±0.737 nmol/g FW.The selenium contents in midgut were 0.100±0.075,4.566±0.505 and 16.036±1.69 nmol/g FW.The selenium content of silk gland was 0.142±0.053,4.266±0.283 and 17.813±0.334nmol/g FW.The selenium contents in epidermis were 0.315±0.026,9.647±0.447 and16.996±0.296 nmol/g FW.The selenium contents of fat body were 0.042±0.073,4.372±0.221 and 12.735±0.543 nmol/g FW.The selenium content of Malpighian tube was0.490±0.065,8.646±0.743 and 13.142±1.038 nmol/g FW,which indicated that the sperm and head of silkworm had the strongest selenium enrichment ability compared with other tissues and organs.Therefore,the results suggested that selenium might regulate the physiological status of silkworm through affecting the reproductive system and neural development.In addition,the selenium enrichment ability of female pupae is stronger than that of male pupae.3.Selenium supplementation had different effects on the expression of selenophosphin-related genes in different tissues.The expression of 9 protein-related genes in different tissues of silkworm was determined by real-time fluorescence quantitative method.including glutathione peroxidase(GPX),thioredoxin reductase(TRXNRD),iodothyronine deiodinase(DIO),selenophosphatase(SPS2),selenoprotein P(Sel P),methionine sulfoxide reductase A(Msr A),selenoprotein T(Sel T),and selenoprotein W(Sel W).The results showed that when selenium concentration increased,the nine selenophenin-related genes regulated tissues were different.GPX: hemolymph,head,midgut,adipose body and testis.SPS2: in each organization;SOD: head,Malpighian tube and testis;DIO: hemolymph and head;TRXNRD: midgut,Malpighian tube,and testis;Sel W: head,midgut,Malpighian tube and testis;Sle P: Malpighian tube and testis;Msr A:head,Malpighian tube,and testis;Prx: head,Malpighian tube and testis.4.The effect of GPX gene knocked down and the effect of selenium supplementation.After GPX gene knocked down 72 hours,the body weight of silkworm decreased significantly,but selenium supplementation had little effect on the influence of body weight knocked down.After GPX gene was knocked down,the total cocoon quantity,cocoon layer quantity and cocoon layer percentage decreased significantly.After add Se the total cocoon quantity,cocoon layer quantity and cocoon layer percentage recovered obviously.After GPX gene knocked down,the number of oviposition and oviposition were significantly reduced,after add selenium supplementation the body weight can be improved.In the interference experiment of GPX gene,the interference effect of the head was the best,followed by testis,Malpighian tube and adipose body.The results of three aspects: physiological indexes,physical and chemical properties and gene level of silkworm growth and development were systematically explained the molecular mechanism of Selenium supplementation in regulating growth,development and reproduction of Bombyx.The results provided the reference value for the study of physiological regulation function,and laid a solid theoretical basis for silkworm new nutrient element selenium development. |
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