Map-based Cloning Of Rice Yellow-green Leaf Gene YGL53 And Its Effects On Carbon And Nitrogen Metabolism Related Characteristics

Leaves are the main place for rice photosynthesis,and the yield of rice mainly depends on the photosynthetic capacity.Improving the photosynthetic efficiency of the population is one of the important goals of rice breeding.In this study,the seeds of Shennong 9816（SN9816）were treated with ⁶⁰Co-γradiation,and the mutant materials were screened by agronomic trait investigation to obtain a stable genetic yellow-green leaf mutant（ygl53）.The whole plant of mutant ygl53 appeared yellow-green throughout the growth period.In this study,the molecular mechanism of the formation of yellow-green leaf mutant ygl53 was elucidated by using techniques and methods related to cell biology,crop physiology,biochemistry and molecular biology.The main results are as follows:1.Chlorophyll synthesis and chloroplast development of mutant ygl53Compared with WT,Magnesium chelatase（Mg CH,E.C.6.6.1.1） activity of mutant ygl53was reduced,resulting in impaired synthesis of Mg-protoporphyrin IX（Mg-proto IX）and the decrease of the chlorophyll（Chl）content,so the leaves of the mutant ygl53 are obviously yellow-green.The chloroplast ultrastructure was observed,and it was found that the thylakoid lamellae of mutant ygl53 were clear,neatly arranged,and rich in starch granules.The thickness and number of lamellae were not significantly different from those of wild type SN9816,indicating that the decrease in Chl content of mutant ygl53 did not affect the chloroplast development.2.Photosynthesis of mutant ygl53Compared with WT,the quantum yield of mutant ygl53 for heat dissipation is reduced by13.25%,the quantum yield for electron transfer is increased by 25.00%,the superoxide anion production rate and H₂O₂ content were reduced,and the carboxylation activity of ribulose-1,5-bisphosphate carboxylase/oxygenase（Ru Bis CO,EC 4.1.1.39）in mutant ygl53 increased.Analysis of photosynthetic gas exchange parameters showed that the net CO₂ assimilation rate of the mutant ygl53 was increased by 24.60%.The mutant ygl53 reduced the content of light-harvesting pigment molecules,reduced the excessive absorption of light energy,reduced the stress of reactive oxygen species（ROS）,increased the efficiency of electron transfer and enhanced the photosynthetic CO₂ assimilation ability.The results of population light transmittance showed that the transmittance at 40 cm and60 cm below the canopy of mutant ygl53 was significantly higher than that of WT by 17.36%and 32.03%,and the leaf senescence under the mutant ygl53 canopy was slower.The results showed that the color of the leaves of the mutant ygl53 was lighter and the light transmittance was easier,which improved the light transmission rate of the population.The leaves in the lower canopy were above the light compensation point,improving the photosynthesis of the population and delaying the leaf senescence process.3.Yield and yield components of mutant ygl53There was no significant difference in dry matter accumulation and distribution ratio between wild type SN9816 and mutant ygl53.However,compared with the wild type,the sheath of mutant ygl53 contributed more to export and transport.The leaf transport of wild type was mainly concentrated from milk stage to dough stage,while the leaf transport of mutant ygl53from milk stage to maturity was all transported.The main yield traits of wild type and mutant ygl53 were not significantly different.Because the mutant ygl53 had lower nitrogen accumulation and higher photosynthetic assimilation rate,its photosynthetic nitrogen use efficiency is increased.Although nitrogen uptake efficiency of the mutant ygl53 was reduced,the higher photosynthetic nitrogen use efficiency helped it maintain its yield without loss.4.The effect of photorespiration on carbon metabolism and nitrogen metabolismThe mutant ygl53 had higher RuBisCO oxidation activity and higher photorespiration rate,and its Glutamate synthase（GOGAT）and Glutamine synthetase（GS）activities were significantly higher than those of the wild type.The nitrogen accumulation of mutant ygl53decreased,and the C/N ratio increased.The increase of photorespiration consumed more carboxylated products to effectively alleviate the continuous increase of C/N ratio.NH₃ andα-Ketoglutaric acid（2-OG）produced by photorespiration can be reassimilated into glutamic acid（Glu）and glutamine（Gln）to supplement nitrogen metabolism through GS/GOGAT.This indicates that photorespiration plays an important role in maintaining the steady state of plant carbon and nitrogen metabolism.5.Map-based cloning and functional verification of YGL53YGL53 was located in the 40.99kb region of chromosome 3 by map-based cloning,containing 7 open reading frames（ORF）.Sequence analysis showed that a single base T at1805th of the 4th ORF was replaced by A,resulting in the substitution of Val-602 with an Asp residue.Gene function complementation and gene editing experiments demonstrated that LOC＿Os03g59640 is the YGL53 gene that controls the yellow-green leaf phenotype,which encodes the D subunit of Mg CH.