| We transferred the anti-trxs gene into wheat cultivar 895004 in order to abtain a transgenic line and tested it by PCR.In addtional,we tested the ear sprouting and seeds germination with positive lines to select lines which have resistance to pre-harvest sprouting.We researched the changes of thioredoxin h activities,its related enzyme activities and carbon metabolism,protein content,respiratory rate of transgenic lines and non-transgenic lines in the process of forming and afterripening and germinating of seeds respectively,the main results were as following:1 The anti-trxs gene was inherited steadily into the T5,T6 transgenic linesWe used PCR test to validate the transgenic plants in T5,T6 lines.The results showed that 10 positive lines among 13 transgenic lines indicated that anti-trxs gene had been inherited steadily into the transgenic lines.2 The transgenic lines had the significant resistance on the ear sprouting and seeds germinationWe choose positive lines to do ear sprouting and seeds germination experiment, the transgenic lines 5-3,38-1,38-3,79-1,97 had the pre-harvest sprouting resistance, but 64-4 had not.The resistant lines were more significant than non-transgenic lines in sprouting time,sprouting rate,sprouting degree and seeds parameter 20 days after maturity,the capacity of seeds germination in positive lines recovered and no significant difference was found among them.The correlation analysis indicated that significant correlations existed between ear sprouting parameter seeds germination parameter.3 The physiological and biochemical characteristics of the transgenic lines3.1 The activities of thioredoxin h and the amylase,protease in the transgenic lines were decreased We had determined the thioredoxin h redox activities and its related enzyme activities in the pure transgenic lines and non- transgenic line.The results showed that the trxh activity in the transgenic lines was lower than that in the non-transgenic line in the process of seeds forming and afterripening,compared with non-transgenic the trxh activity degraded averagely by 42.2%(P＜0.01)at the period of 30d after anthesis to 10d after ripeness.The trxh activity lessening leaded to amylase and protease activities reducing,compared with non-transgenic seeds,the a-amylase activity in transgenic seeds decressed averagely by 50.2%(P＜0.01)in the process of seeds forming and afterripening;theβ-amylase activity decressed averagely by19.3%during 30 days after anthesis to ripeness;the DBE activity degraded averagely by 27.4%(P＜0.05)from ripeness to 10 days after maturity;the protease activity difference appeared after ripeness and decressed averagely by 37.4%(P＜0.05).In the germination the activity of amylase and protease increased,and they were lower in the transgenic lines than that in the non-transgenic line.Theα-amylase activity in transgenic seeds were restrained obviously during all the seeds germination;but theβ-amylase activity started to produce an effect mainly after 48 hours;the DBE activity effected mainly before 48 hours,the protease activity played a role at 12 hours after germination.When we transferred anti-trxs gene into wheat,the trxh activity reduced in wheat,thioredoxin(Trx)plays an important role in regulating activity of amylase and protease by deoxidizingS-S to S-H in wheat plant to inhibit pre-harvest sprouting.3.2 The starch constituent contents were increased and the dissolvable sugar content were degraded in the transgenic lines The result of starch content in the transgenic seeds and non- transgenic seeds showed that:the content of starch amylose and amylopectin in the transgenic seeds was higher than non- transgenic seeds, amylopectin /amylose was at the same level and had not difference.The starch degradation speed was obviously slower than non- transgenic seeds.The difference of the content of dissolved sugar between the transgenic seeds and non-transgenic seeds was obvious during the 35 days after anthesis to 20 days after maturity.During the seeds germination,the content of dissolved sugar increased gradually,the upstroke velocity in the non-transgenic seeds and transgenic seeds were 0.22 and 0.07 respectively,which indicated that carbon metabolism was slow and decreased the supply of carbon in order to inhibit the pre-harvest sprouting.3.3 The albumin and glutelin content in the transgenie lines were increased,the storage protein were degraded slowly in seeds germination In the process of seeds forming and afterripening,the content of albumin and globulin decreased,but the content of gliadin and glutelin increased.The content of albumin and glutelin in the transgenic seeds increased 20.6%and 14.6%respectively.During the seeds germination,the content of gliadin and glutelin decreased with a straight line as germination time extended and their decreasing rate in the transgenic seeds were 55% to contrast.This indicated that the anti-trxs expression restrained protease activities to hamper storage protein degradation.3.4 The respiratory rate of transgenie lines were cut down In the period of grain maturing and after ripening,the respiratory rate depressed gradually and it reached lowest at maturity,which showed the curve of peak variation at the period of after-ripening.The respiratory rate in the transgenic seeds was lower than that in the non-transgenic seeds,and it decreased averagely 42.3%from 20d after anthesis to 30d after maturity.The contrast appeared a climacteric at 10d after ripening,but the tansgenic lines didn't have.The coefficient of variability in the transgenic seeds and non-transgenic seeds were 65.2%and 51.7%from mature to 30d after mature.During seeds germination,the respiratory rate showed zoom tendency and its upstroke velocity of the transgenic lines was 44.6%than that of non-transgenic lines.Compared to non-transgenic lines,the respiratory rate in the transgenic seeds decreased 34.9% averagely.Which indicated that the expression of the anti-trxs gene had an inhibitory action on respiratory.
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