The Effect Of Polyamine On The Growing Development And Cloning The ODC Gene Fragment Of Helicoverpa Armigera

Polyamines,such as putrescine,spermidine and spermine,are ubiquitous physiological cations have two or more than two amines.They occur in almost all living species,both prokaryotes and eukaryotes.Polyamines are essential for the synthesization of large biologic molecule,cell growth and differentiation.They can also regulate the physiological process of insect such as growth and development et al. Ornithine decarboxylase(ODC,EC4.1.1.17) is a key enzyme in polyamine biosynthesis and decarboxylates L-ornithine to putrescine.In animals,the regulation of polyamine levels by ODC plays an important role in the control of growth, proliferation and differentiation of cells.The activity of ODC rises markedly in response to various growth and differentiation stimuli.On the other hand,ODC is under negative feedback regulation exerted by polyamines.Whereas the regulation mechanism is unclear because of the complexity of the physiological process of insect and the metabolism of polyamines,also we have not seen the reporting between the ODC and the insect.So,studying the effect on growth and development,the synthesize of yolk proteins,contents of polyamines in Helicoverpa armigera and cloning the ODC gene are essential for understanding the regulation function of polyamines on the physiological process of insect.In this study,on the basis of studying the function of Put on the growth and development and the content changing of Helicoverpa armigera,we Analysed the mechanism of action of polyamines to the insect,by cloning the ODC gene fragment,we analysed the structure characteristics, inquiring the functioning of the ODC.The main results as follows:1 The effect of putrescine on the development and growth of the Helicoverpa armigeraTo investigate the effect on the developmental phase of laraval,the weight of chrysails and the abnormality of chrysails and eclosion of the Helicoverpa armigera, we use four different concentration of putrescine and 0.05%concentration of putrescine feeding the Helicoverpa armigera.The results indicate:After treated with four different concentration of putrescine,the developmental phase of laraval is prolonged;and the developmental phase which treated with 0.01%Put is the longest, reach 17.72±0.04d,have very significant difference compared with the control,with the increasing concentration of putrescine,the developmental phase of laraval is gradual declined,the development duration of chrysalis have great change,and the development duration of chrysails treated with 0.1%and 0.05%Put reach very significant difference compared with the group treated with 0.01%Put,the group treated with 0.5%Put have significant difference compared with 0.1%Put,but the group treated with 0.01%Put,0.5%Put have no significant difference with control; the weight of chrysails have different degree increasing compared with control,the weight of chrysails treated with 0.01%Put has the maximum weight,reach 0.2806±0.0032g,but the other three group have no significant difference compared with the control;the abnormality of chrysails all increasing significantly after treated with four different concentration of putrescine,and the group treated with 0.05%Put and 0.1%Put have a higher abnormality compared with the group treated with 0.01%Put and 0.5%Put;with the increasing of the concentration of the putrescine,the abnormality of the eclosion is declined.After continued treated with 0.05% concentration of putrescine,the developmental phase of laraval and the weight of chrysails have no significantly difference compared with control.2 The effect of putrescine on the oviposition of the Helicoverpa armigeraThe results indicate:the average oviposition number have no significant difference compared with the control after treated with four different concentration of putrescine, but the average oviposition number has the maximum treated with 0.01%put,reaching 1001.9±63.1 egg/female,and the 0.5%put has the minimum number,indicate the low concentration of putrescine(0.01%put) can increase the average oviposition number, and the high concentration of putrescine depressed.After continued treated with 0.05%concentration of putrescine,the total oviposition number of the first generation has no obviously difference compared with the control,but the total oviposition number of the second generation significantly declined,only have 498 egg/female, has significant difference compared with the control,the total oviposition number of the third and the fouth generation are return,but have no significant difference compared with control.3 The effect of putrescine on the content changing of Polyamine of the Helicoverpa armigeraThe results indicate that the content of the putrescine and spermidine in laraval of the Helicoverpa armigera has changed significantly,but the spermine has low level at all time.After treated with low concentration of putrescine,the content of polyamine have changed,but the high concentration of putrescine has no effect on the content of polyamine.After continued treated with 0.05%put,the content of polyamine have greatly changed,and the content of polyamine of the fouth generation has significantly difference compared with the control and the other three generation.4 The effect on the synthesize of yolk proteins of Helicoverpa armigera of polyaminesThe results of SDS-PAGE showed that the vitellin has two chains,the molecular mass of the heavy chain is 165KD,and the molecular mass of the light chain is 50KD.The results showed that:the synthesis of the yolk proteins were syimulated when the group treated with 0.1%Put and 0.05%Put,and in the most of the periods of development,the content of the vitellin was obviously higher compares with control; but the synthesize was restrained when treated with 0.01%Put and 0.5%Put,and the content of the vitellin was lower compared with the control.After eclosion 36h,the contents of the yolk proteins slow declined when treated with low concentration of putrescine(0.01%put),but the contents of the yolk proteins rapidly declined when treated with high concentration of putrescine.5 The ODC gene fragment cloning of the Helicoverpa armigeraOn the base of the known conserved sequence of the Ornithine decarboxylase gene, we designed one pair of ODC degenerate primer,proceeding RT-PCR,achieving a objective band which is similar to the objective band,and achieving a fragment of the Omithine decarboxylase gene of Helicoverpa armigera.