| Nitrogen(N)is one of the most important nutrients in potato growth.N fertilizer plays a vital role in the formation of potato tuber yield and quality.The purposes of this research were to investigate the physiological and molecular regulation mechanism of the effect of N fertilizer application rate on the accumulation of potato tuber starch.Two potato varieties with similar growth period but different starch content(high starch variety Kexin 22 and low starch variety Kexin 19),which were assigned in main plot,and five N application rates(0,75,150,225 and 300 kg/ha)assighed in sub-plot in a split plot design,were studied for their effects on the agronomic characters,starch accumulation,starch accumulation rate,starch synthesis key enzyme activity,related gene expression,and their relationship among the traits tested.Also,the effects of N application rate on the microstructure and quantity of potato starch granules were investigated using scanning electron microscope and transmission electron microscope.Results showed that:(1)The accumulation of amylose,amylopectin and total starch in Kexin 22 showed a ranking of N75>N150>N225>N300>N0 at both early and middle growth stages,and N150>N225>N300>N75>N0 at the late growth stage.The accumulation of amylose,amylopectin and total starch in Kexin 19 showed a pattern of N75>N0>N150>N225>N300 in the early growth period,and N75>N150>N225>N300>N0 in the middle growth period.For amylose accumulation at the late growth stage,Kexin 19 showed a ranking of N300>N225>N150>N75>N0,however,the accumulation of amylopectin and total starch were in the rank of N225>N300>N150>N0 in the late growth period.Compared with Kexin 22,Kexin 19 was very sensitive to N fertilizer,and it is a nitrogen-loving variety.In comparison of the two varieties,amylose,amylopectin,and total starch accumulation of Kexin 22 were significantly higher than those of Kexin 19 during the whole growth period.(2)The accumulation rate of amylose,amylopectin,and total starch in Kexin 22 showed a ranking of N75>N150>N225>N300>N0 at the early growth stage,N150>N225>N300>N75>N0 at the middle growth stage,and N300>N225>N150>N75>N0 at the late growth stage.The appearance time of peak value of amylose,amylopectin and total starch accumulation rate was not changed under different N fertilizer rates,all being at 65 days after emergence.For Kexin 19,the accumulation rate of amylose,amylopectin,and total starch showed a pattern of N75>N0>N150>N225>N300 at the early stage of growth.In the middle period of growth,the accumulation rate of amylose showed a ranking of N150>N225>N300>N75>N0,amylopectin showed N150>N75>N225>N300>N0,and total starch showed N150>N225>N75>N300>N0.At the late growth stage,amylose,amylopectin,and total starch accumulation rates all showed a ranking of N300>N225>N150>N75>N0.The peak times of the accumulation rate of amylose,amylopectin,and total starch at the low nitrogen rate were obviously advanced,while the peak times of the accumulation rate of amylose,amylopectin and total starch were obviously delayed at the high nitrogen rate.In comparison of the two varieties,at the early growth stage,no significant differences were found for amylose,amylopectin,and total starch accumulation rate between Kexin 22 and Kexin 19;at the middle growth stage,amylose,amylopectin,and total starch accumulation rate of Kexin 22 were significantly higher than those of Kexin 19;and at the late growth stage,amylopectin and total starch accumulation rate of Kexin 22 were significantly higher than those of Kexin 19.(3)The activities of AGPP,GBSS,SSS,and SBE in Kexin 22 showed a trend of N75>N150>N225 >N300>N0 at the early growth stage,and N150>N225>N300>N75>N0 at the middle growth stage,and N300>N225>N150>N75>N0 at the late growth stage.The peak values of the activity of these four enzymes all appeared on 65 days after emergence under different nitrogen rates.For Kexin 19,the enzyme activity of AGPP,GBSS,SSS,and SBE showed a trend of N75>N0>N150>N225>N300 at early growth stage,N225>N300>N150>N75>N0 at middle growth stage,and N300>N225>N150>N75 >N0 at late growth stage.At low nitrogen rate,the appearance times of the peak activity of AGPP,GBSS,SSS,and SBE appeared obviously earlier,however,at the high nitrogen rate,the times of the peak activity of these enzymes were obviously delayed.In comparison of these two varieties,the enzyme activities of AGPP,GBSS,SSS,and SBE of high starch variety Kexin 22 were significantly higher than those of the low starch variety Kexin 19.At different nitrogen rates,the accumulation rate of amylose was positively correlated to the activity of AGPP and GBSS,and the accumulation rate of amylopectin was positively correlated to the activity of AGPP,SSS,and SBE enzymes.The accumulation rate of total starch was positively correlated to the activity of AGPP,GBSS,SSS,and SBE enzymes.The change trend of starch accumulation rate with N application rate was generally in accordance with the change trend of the activity of the corresponding starch synthase.(4)In comparison with the variety of Kexin 19,gene expression of AGPP-L,GBSSI,SBEI,and SSIII of Kexin 22 increased obviously,whereas no obvious difference was observed for gene expression of AGPP-S and SSII.Different N application rates significantly impacted peak values of gene expression of AGPP-L,GBSSI,SBEI,and SSIII of Kexin 22,whereas it did not greatly influence the peak value appearance time of gene expression.For Kexin 22,gene expression of AGPP-L,GBSSI,SBEI,and SSIII mostly showed a ranking of N75>N150>N225>N300>N0 at the early growth stage;at the middle growth stage,mostly showed N150>N225>N300>N75>N0;and at the late growth stage,all showed N300>N225>N150>N75>N0.Under different N fertilizer application rates,the appearance times of peak value of these gene expressions mostly appeared on 65 days after emergence.Various N fertilizer application rates significantly influenced the levels of gene expression and peak times of gene expression of AGPP-L,GBSSI,SBEI,and SSIII of Kexin 19.As for the levels of these gene expression,at the early growth stage,they all showed a ranking of N75>N0>N150>N225>N300;at the middle growth stage,they mostly showed N150> N225>N300>N75>N0,and in the late growth period,they all showed N300>N225>N150>N75>N0.Under low N fertilizer condition,the appearance time of peak value of gene expression was advanced;whereas under high N fertilizer application condition,an obvious delay was found for the appearance time of peak value of gene expression.The expression of key enzyme genes in starch synthesis is similar to the trend of corresponding starch synthetase activity with increase in nitrogen application.(5)At the early stage of tuberization,with the increase of N application rate,the size of tuber parenchyma cells in Kexin 22 and Kexin 19 became smaller.The number of large-sized starch granules decreased obviously,while the number of small-sized starch granules increased dramatically.For Kexin 22,when the rate of nitrogen applied was in the range of 0-225 kg/ha,in the tuber maturing period,the size of parenchyma cells in the tuber increased obviously,and the number of large-sized and small-sized starch granules also increased obviously with the increase of nitrogen application.When the rate of nitrogen application was in the range of 225-300 kg/ha,with the increase of nitrogen application,the size of the parenchyma cells in the tubers became smaller,and large-sized starch granule and total starch granule number apparently decreased.For Kexin 19,when the rate of nitrogen applied was in the range of 0-225 kg/ha,in the tuber maturing period with the increase of nitrogen application,the size of the tuber parenchyma cells increased obviousely.The number of large-sized starch granule increased obviously,while the number of small-sized starch granules decreased remarkably.When nitrogen application rate was in the range of 225-300 kg/ha,with the increase of nitrogen application,the size of parenchyma cells did not change much,but the number of large-sized starch granules greatly decreased,however,the number of small-sized starch granules dramatically increased.Compared with the growth period,the size of the parenchyma cells and starch granules of the two varieties obviously increased from the early tuber initiation to maturity stage.Compared with the two varieties,the sizes of the tuber parenchyma cells and starch granules of Kexin 22 were obviously larger than those of kexin 19.Also,the number of large-sized starch granules of the tuber of Kexin 22 was greater than that of Kexin 19.(6)There are three ways of multiplication of the amyloid in the tubers of both Kexin 22 and Kexin 19,including multinuclear,constriction,and budding.At early tuber initiation stage,with the increase of nitrogen application,the multiplication rate of starch amyloids in the two varieties increased first and then decreased,with the rate of the multiplication being the highest when the nitrogen application rate was at 75 kg/ha.At tuber maturity stage,with the increase of nitrogen application,the multiplication rate of starch amyloids of the two varieties also increased first and then decreased.When the rate of nitrogen application was at 150 kg/ha,the multiplication rate of starch amyloids of Kexin 22 was the highest,while when nitrogen application rate was at 225 kg/ha,the multiplication rate of Kexin 19 was the highest.Compared with the two varieties,the rate of starch amyloid multiplication of Kexin 22 was obviously higher than that of Kexin 19.In comparison with the growth stage,the rate of starch amyloid multiplication was obviously higher at early tuber initiation stage than that at tuber maturity stage. |
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