| Rice FOT is the “Flower Opening Time” after heading in one day.It usually refers to the peak flowering opening time.In China,hybrid rice has made great contributions to food security.However,the missing FOT of sterile lines and restorer lines is one of key factors restricting the yield of hybrid rice seed production.Furthermore,for indica and japonica interspecies hybrid rice with greater heterosis,the contradiction is more prominent,which limits the application and promotion of many excellent rice germplasm resources.Therefore,searching for genetic and artificial FOT improvement methods to synchronize the sterile line with the restoring line has always been a concern of rice breeders.At present,there are few studies on the molecular mechanism of rice FOT regulation,and it is urgent to further study.In the previous study,we screened and identified an early flower opening time mutant y107 in a library of ethyl methane sulfonate(EMS)mutagenic mutants in the background of Nipponbare.Genetic analysis showed that this early flower opening time trait was controlled by a pair of recessive nuclear genes.Then,the candidate gene Y107 was cloned using Mut Map,and further research was carried out,and the following experimental results were obtained.1、The correctness of the cloned gene Y107 was confirmed by transgene complementation,RNAi interference and CRISPR / Cas9 knockout.The flower opening time trait investigation found that the flower opening times of RNAi interference plants and CRISPR/Cas9 knockout plants were the same as the flower opening time of the y107 mutant,and the complementary lines were the same as those of the wild type Nipponbare.2、Promoter GUS staining showed,in spikelets,the Y107 gene was mainly expressed in lobe slices.In addition,Y107 was also highly expressed in the root tip,leaf and stem vascular.Quantitative RT-PCR results showed that Y107 expressed higher in the lower half of the entire spikelet,indicating that the Y107 gene might play an important role in the regulation of lodicule.3、The detection of the expression of the Y107 gene in unopened florets revealed that the expression of the Y107 gene gradually increased before the opening of florets and gradually decreased after the closure of florets,indicating that the expression of Y107 in florets has a certain rhythm.4、The key gene of the jasmonic acid pathway Os JAR1 was knocked out by CRISPR / Cas9 against the background of Nipponbare.The mutant osjar1 exhibited later FOT and unclosed glumes;the double mutant y107/osjar1 exhibited the same FOT as osjar1,indicating that the Os JAR1 may act downstream of the Y107 gene.5、Using CRISPR/Cas9 to knock out Y107 in wild type Yixiang 1B,the FOT1 of Yixiang 1B knockout mutant was about an hour earlier than that of wild type Yixiang 1B.Accordingly,Y107 could control FOT in both indica and japonica rice,suggesting the potential of Y107 in application.6、Jasmonic acid plays an important role in the regulation of rice FOT.The FOT of y107 after methyl jasmonate treatment was the same as that of wild type,suggesting that Y107 may be related to the jasmonic acid pathway.Therefore,we detected the expression of jasmonic acid related genes.Compared with the wild-type,the expression levels of the jasmonic acid related genes Os JAR1,Os AOS1,Os DAD1,Os OPR7,Os JAZ1 and Os MYC2 in Yixiang 1B knockout mutant reached the peak in advance after 11 o’clock,suggesting that Y107 may regulate rice FOT through the jasmonic acid pathway. |
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