Phylogenetic Analyses Of NDH Complex Related Nuclear-encodec Genes In Maize And Study On Salt Tolerance Photosynthetic Mechanism Of Ndhl Subunit-overexpressed Rice Lines

With the increase of saline-alkali land,salt stress has become an important abiotic stress factor restricting crop yield.Exploring the physiological mechanism of crop salt tolerance to improve food production and quality is the constant pursuit of human goals.NDH complex mediated cyclic electron transport around PSI(CET-PSI)is a very important protective mechanism for plants to adapt to a variety of natural environmental stresses in the high-plant.Due to the requirement of extra ATP for driving the CO2-concentrating mechanism in C4 plants,the ability of CET in C4 plants is stronger than in C3 plants.The difference of NDH expression between C4 plant and C3 plant was the main factor that caused the capacity difference of CET.Among the three major food crops,maize and rice belong to C4 and C3 plants,respectively.In this study,we identified about related nuclear-encoded genes of NDH complex from the whole genome of maize based on bioinformatics.And the phylogenetic analysis of the identified genes was carried out based on the gene family.Screening genes that may function in C3 and C4 plants,and the ZmNdhLl gene screening from gene family experienced positive selection was selected to study the transgene overexpression in rice.We compared and analyzed the characteristices on photo synthetic physiology of two transgenic lines and wild type plants under salt stress.The main results are as follows:1.Searching the reference genomes of the Zea mays,arabidopsis thaliana,Oryza sativa L.and Sorghum based on the 17 neclear subunits of NDH complex with their protein domains.We identified 27 maize genes,22 arabidopsis genes,23 rice gene and 23 sorghum genes.The phylogenetic tree of NDH nuclear gene was divided into three categories according to gene repeats and tree map.A type:there are 8 phylogenetic trees that is a single cluster built by four species and which are single copy gene;B type:there are 5 evolution trees that is a single cluster built by four species,but there are two copies in maize or sorghum.C type:There are four phylogenetic trees with two or more homologous gene clusters.Two gene families NdhL and PnsB2 were screened by positive selection,which experienced positive selection in maize.2.Real-time quantitative analysis of two gene families NdhL and PnsB2 in maize showed that ZmNdhLl had the highest expression level.ZmNdhLl was highly expressed in leaves and induced by abiotic stress,such as salt,drought and low temperature.The full length cDNA of the ZmNdhLl gene was cloned from the maize leaves of ZM395 which has a stronger CET activity,and its open reading frame(ORF)was 588bp,encoding 195 amino acids.The ZmNdhLl overexpression vector was constructed and transformed into rice by agrobacterium tumefaciens.Then twenty-four transgenic plants were screened by hygromycin resistance identification,PCR and RT-PCR.The expression of OE3 and OE13 was the highest in transgenic plants and was used for subsequent studies.3.Determination of the degree of curl and the chlorophyll fluorescence kinetics curves of transgenic and wild type plant leaves,the results showed that there was no significant difference between overexpression lines and wild type under normal conditions.Compare to the wild type in the condition of 200mM salt treatment,the results of the overexpression lines showed that:The blade curl was lower;The increase of chlorophyll fluorescence instantaneously after exposure to light was higher;The decrease of maximum photochemical efficiency of PSI(Y(?)),the maximum photochemical efficiency of PSII(Y(?)),the electron transport rate of PSI(ETRI),the electron transport rate of PSII(ETRII)are smaller;The J point of chlorophyll fluorescence OJIP curve is down,the peak of K-band is smaller;The decline range of I1 and I2 point in delayed fluorescence induction curve is smaller.The results showed that the salt tolerance of overexpression lines was better than that of wild type.The overexpression of the ZmNdhLl gene enhances the CET activity of the plant,making the oxygen-evolving complex(OEC)of the photo synthetic electron transport chain and the activity of the electrons transferred from the original quinone receptor QA are smaller.Thereby reducing the damage of PSI and PSII by the salt stress.