| As one of the famous “Four Huaiqing Chinese medicines”,Dioscorea opposita Thunb.is not only delicious but also medicative,and can be a good nutrition and health product.The virus-free microtuber is an important source of high quality seedlings of Dioscorea opposita Thunb.and it can solve the problem of virus diseases and quality degradation caused by long-term vegetative propagation.On the basis of previous research work,in order to ensure the virus-free microtuber produced in different periods germinate uniformly and efficiently,we carried out a systematic study on the germination of the virus-free microtuber and explored the germination conditions of the virus-free microtuber mainly from plant growth regulators,volume size,storage temperature and period,etc.In addition,we also explored the mechanism of the germination of virus-free microtuber from the aspects of morphology and anatomy,physiological mechanism and endogenous hormones.The results were as follows:1.The effects of different concentrations of plant growth regulators(NAA,6-BA,KT)on the germination of the virus-free microtuber were great.The optimal combination of plant growth regulators for the germination of the virus-free microtuber was NAA 0.02 mg·L-1and 6-BA 1 mg·L-1 and at this time,the germination rate of the virus-free microtuber was the highest,which was 77.78 %.2.The germination rate of the grade III virus-free microtuber whose diameter was between 0.4 cm and 0.6 cm was higher than the grade?virus-free microtuber whose diameter was greater than 0.8 cm but lower than the grade II virus-free microtuber whose diameter was between 0.6 cm and 0.8 cm no matter in vitro and out of vitro.And the germination rate of the grade II virus-free microtuber could reach 84.49% in vitro and 83.47 % out of vitro.It could be seen that the virus-free microtuber whose volume size was too large or too small was hard to germinate.3.The effects of different storage temperatures(4 ?and 25 ?)on the germination of the virus-free microtuber were great.The virus-free microtuber could germinate more easily and the germination rate of the virus-free microtuber was higher under 4 ? condition than that of under 25 ? condition no matter in vitro and out of vitro.In addition,the germination rate of the virus-free microtuber reached the highest(in vitro 73.40% and out of vitro 78.56%)when the the virus-free microtuber was stored at 4 ?for 6W and changed little when the virus-free microtuber was stored between 6 w and 12 w.4.The internal morphology of the virus-free microtuber and the distribution of starch and protein in the process of the germination had a great change.The outermost of the virus-free microtuber was chapped corky epidermis and within the epidermis was periderm within which was parenchyma.At this time,there were lots of starch grains and protein distributing in central parenchyma cells.When the dormancy of the virus-free microtuber gradually released,the apical meristem cells began to divide and the bud began to sprout.Along with the complete breaking of the dormancy of the virus-free microtuber,the division rate of apical meristem cells was accelerating and the growth cone became a hemisphere.Finally,the budlet appeared.In this process,the content of starch grains and protein continuously reduced.5.In the harvesting stage,seeding stage,hair-root stage,germination stage I,germination stage II and germination stage III,the main organic matters and related enzyme activities and endogenous hormones in the apical bud tissue and other tissue of the virus-free microtuber changed a lot.(1)Changes of main organic mattersDuring the whole process of germination,the starch content in the apical bud tissue and other tissue of the virus-free microtuber dropped.The content of soluble sugar,sucrose,fructose and glucose generally showed a trend of increasing first and then decreasing.The content of total phenol showed a tendency to decrease first and then increase.And in the seeding stage,the content of total phenol decreased to the lowest value.In the apical bud tissue of the virus-free microtuber,the content of protein decreased firstly and then rose and decreased at last.However,the content of protein in other tissue of the virus-free microtuber dropped continuously.(2)Changes of related enzyme activitiesDuring the whole process of germination,the alpha-amylase and starch phosphorylase worked together to complete the decomposition of starch.The activity of alpha-amylase reached the maximum value in the germination stage II and the activity of starch phosphorylase reached the maximum value in the hair-root stage.The invertase and sucrose synthetase worked together to complete the decomposition of sucrose and in the hair-root stage,the activity of invertase and the decomposition activity of sucrose synthetase appeared maximum value.The activity of polyphenol oxidase directly affected the content of total phenol and when the content of phenolic compounds reached the lowest value in the seeding stage,the activity of polyphenol oxidase reached the maximum value.In addition,by adjusting the activity of SOD,POD and CAT,the antioxidant enzyme system kept the inner membrane system of the virus-free microtuber in a stable level.(3)Changes of endogenous hormonesThe content of GA,IAA,ZR both in the apical bud tissue and other tissue of the virus-free microtuber was very low and the content of ABA was very high in these two tissues when the virus-free microtuber was in the harvesting stage.In the seeding and hair-root stage,the content of GA,IAA and ZR increased but the content of ABA decreased in the apical bud tissue of the virus-free microtuber.In the germination stage I,the content of GA and ABA increased but the content of IAA and ZR decreased slightly in the apical bud tissue of the virus-free microtuber.In the germination stage II,the content of GA,ZR and ABA entered a downward trend and the content of IAA slightly raised in the apical bud tissue of the virus-free microtuber.In other tissue of the virus-free microtuber,the content of GA did not change obviously and the content of ABA and IAA increased along with the content of ZR decreasing.In addition to the content of IAA in the apical bud tissue showed a rising trend and the content of ABA in other tissue showed a decreasing trend,the content of other hormones whether in the apical bud tissue or other tissue did not obviously change in the germination stage III.In a word,GA,IAA and ZR promoted the dormancy release and germination of the virus-free microtuber while ABA showed a inhibitory effect on the dormancy release and germination.In addition,the content of JA showed a decreasing tendency first and then increased during the whole process.Therefore,we thought that JA may play an inhibitory role in the process of dormancy release and play a promoting role in the later germination of the bud for the virus-free microtuber. |
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