| Waxy maize originated in China,is one of healthy food deeply loved by the people of our country because of its sweet taste and good palatability.Waxy maize plays an important role in textile,papermaking,brewing,feed and other industries as an important raw material because most starch in waxy maize endosperm is amylopectin,has high economic value.However,the nutritional value of waxy maize is not complete,especially the lack of essential amino acids-lysine and tryptophan,can not meet the nutritional needs of human consumption and livestock and poultry breeding.Therefore,it is of great significance to increase the content of lysine in waxy maize endosperm to improve the nutritional quality,edible value and commodity value of waxy maize.The introgression of opaque2 mutant gene into waxy maize inbred lines can greatly increase the lysine content in kernels,but at the same time make the kernel into powdery form,which hinders its application in production.In this study,several opaque2-waxy double recessive near isogenic lines(o2w2-NIL)were constructed by introgressing opaque2 gene of high quality protein maize(QPM)containing o2 modifiers into waxy maize inbred lines,and a series of comparative analysis of morphology,quality,sub-microstructure and protein expression level between o2w2-NIL and their receptors and donors were carried out.The influence of opaque2 on different waxy maize was preliminarily determined,which provided the theoretical basis for the improvement of waxy maize using opaque2.The main results of this study are as follows:1.o2w2-NIL were constructed by backcrossing technique and molecular marker-assisted selection.The lysine content in kernels of 5 o2w2-NIL and their donors and receptors were detected.The results showed that lysine content of Zhong97-7/o2o2,Zhong63-2/o2o2,Zhao OP-6/o2o2,300 MP/o2o2 and Zhong501/o2o2 were increased by 59.0%,52.7%,48.5%,46.3% and 61.9%,respectively.2.5 sets of o2w2-NIL were continuously propagated for 3 generations,and their stability was investigated.The results showed that all the 5 o2w2-NIL could be stably inherited,and the Bg-rbg transposon system had no effect on their stability.3.A series of comparative analysis were carried out between o2w2-NIL Zhong97-7/o2o2,Zhong63-2/o2o2,Zhao OP-6/o2o2,300 MP/o2o2 and Zhong501/o2o2 and their donors and receptors,found that there were differences in the contents of water,crude fat,crude protein,crude starch,total amino acids and 17 kinds of amino acids,and the content of amylopectin and non-zein increased significantly.4.The endosperm sub-microstructure of Zhong97-7/o2o2,Zhong63-2/o2o2,300 MP/o2o2 and their donors and receptors was observed by scanning electron microscopy and transmission electron microscopy.The results showed that the shape,size and distribution density of starch granules and protein bodies in endosperm cells were quite different,and the sub-microstructure had some synchronization with kernel phenotype.5.The expression of O2 protein was significantly inhibited by the introgression of o2 mutation gene,and the expression of WX protein was very low in waxy maize.The relative expression level of O2 and WX protein in 10 DAP,14DAP,18 DAP,22DAP and 26 DAP was detected by qRT-PCR.The results showed that the relative expression level of O2 and WX protein increased with time,then decreased after stabilization.6.The O2 transcription factor was proved to have no direct interaction with WX and wx gene by yeast one hybrid technique.It is speculated that O2 and WX may play an indirect interaction by regulating the synthesis of key enzymes in starch synthesis process such as GBSSⅠ. |
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