Editing of rice (Oryza sativa L.) OsMKK3 gene using CRISPR/Cas9 decreases grain length by modulating the expression of photosystem components.

Grain size is one of the most important agronomic traits for grain yield determination in rice. To better understand the proteins that are regulated by the grain size regulatory gene OsMKK3, this gene was knocked out using the CRISPR/Cas9 system, and tandem mass tag (TMT) labeling combined with liquid chromatograph-tandem mass spectrometry analysis was performed to study the regulation of proteins in the panicle. Quantitative proteomic screening revealed a total of 106 differentially expressed proteins (DEPs) via comparison of the OsMKK3 mutant line to the wild-type YexiangB, including 15 and 91 up-regulated and down-regulated DEPs, respectively. Pathway analysis revealed that DEPs were enriched in metabolic pathways, biosynthesis of secondary metabolites, phenylpropanoid biosynthesis, and photosynthesis. Strong interactions were detected among seven down-regulated proteins related to photosystem components in the protein-protein interaction network, and photosynthetic rate was decreased in mutant plants. The results of the liquid chromatography-parallel reaction monitoring/mass spectromery analysis and western blot analysis were consistent with the results of the proteomic analysis, and the results of the quantitative reverse transcription polymerase chain reaction analysis revealed that the expression levels of most candidate genes were consistent with protein levels. Overall, OsMKK3 controls grain size by regulating the protein content in cells. Our findings provide new candidate genes that will aid the study of grain size regulatory mechanisms associated with the mitogen-activated protein kinase (MAPK) signaling pathway.

Development of molecular marker and introgression of Bph3 into elite rice cultivars by marker-assisted selection.

The brown planthopper (BPH) is a serious insect pest of rice and a substantial threat to rice production. Identification of new BPH resistance genes and their transfer into modern rice cultivars are effective breeding approaches to reduce the damage caused by BPH. In this study, we mapped a BPH resistance gene to a 50-kb genomic interval between two InDel markers 4M03980 and 4M04041 on the short arm of chromosome 4 in indica rice cultivar BP60, where the BPH resistance gene was mapped in Rathu Heenati by Liu et al. (2015) and named "Bph3". This region contains two annotated genes Os04g0201900 and Os04g0202300, which encode lectin receptor kinases responsible for BPH resistance. We also developed a molecular marker "MM28T" for Bph3, and introgression Bph3 into susceptible rice restorer lines Guihui582 and Gui7571 by the marker-assisted selection (MAS) approach. The BPH resistance level is significantly enhanced in the Bph3-introgression lines, the resistance scores decrease from 8.2 to 3.6 for Guihui582 and decrease from 8.7 to around 3.8 for Gui7571. Therefore, developing molecular markers for the BPH resistance gene Bph3 and using them for molecular breeding will facilitate the creation of BPH-resistance rice cultivars to reduce damage caused by BPH.

[Geographical populations of brown planthopper in Nanning of South China and in Vietnam: a comparative study].

A comparison was made on the differences of the geographical populations of brown planthopper (BPH), Nilaparvata lugens (Stål), collected from Nanning of South China and from Ha Noi, Ha Tinh, Hue, TP. Ho Chi Minh, and Song Cuu Long of Vietnam. The BPH populations from Nanning, Ha Noi, Ha Tinh and Hue could damage the rice varieties with Bph1 and bph2 genes, whereas the BPH populations from TP. Ho Chi Minh and Song Cuu Long showed stronger virulence to the rice varieties with Bph1, bph2, Bph3 and bph4 genes than the other populations. The survival rates of the nymphs from Nanning, Ha Noi, Ha Tinh and Hue populations on the rice varieties TN1, Mudgo and ASD7 were all above 52%, but those on the rice varieties Rathu Heenati, Babawee and Ptb33 were below 44%. The nymphs of the BPH populations from TP. Ho Chi Minh and Song Cuu Long on the TN1, Mudgo, ASD7, Babawee and Ptb33 had a survival rate of above 50%, but those on Rathu Heenati had a survival rate of below 50%. The nymphal development durations of the BPH populations from Nanning, Ha Noi and Hue were significantly shorter on TN1, Mudgo and ASD7 than on Rathu Heenati, Babawee and Ptb33, but for the BPH population from Ha Tinh, the nymphal development duration on TN1, Mudgo and ASD7, and on Rathu Heenati and Ptb33 had no significant difference. The nymphal development durations of the BPH population from TP. Ho Chi Minh on TN1 and ASD7 were significantly shorter than those on Rathu Heenati, Babawee and Ptb33, but for the BPH population from Song Cuu Long, the nymphal development durations on TN1, Mudgo, ASD7, Rathu Heenati, Babawee and Ptb33 had no significant difference. After infested for 192 h, the nymph density of the BPH populations from Nanning and Ha Noi was significantly lower on Rathu Heenati, Babawee and Ptb33 than on TN1, Mudgo and ASD7, that of the BPH populations from Ha Tinh was significantly lower on Babawee than on TN1, Mudgo, the BPH populations from Hue and TP. Ho Chi Minh had a significantly lower nymph density on Babawee than on TN1, Mudgo, ASD7 and Rathu Heenati, and the BPH population from Song Cuu Long had a significantly lower nymph density on Babawee and Ptb33 than on TN1 and ASD7. The virulence of the BPH population from Nanning was similar with that of the BPH populations from central and northern Vietnam, which was a new proof that the BPH in South China was migrated from the central and northern parts of Vietnam.

[Resistance stability of rice varieties to different biotypes of brown planthopper].

In order to develop an effective analytical and monitoring method in breeding rice varieties with higher resistance stability to brown planthopper (BPH), Tai's method (1971) was employed to estimate the resistance stability of twelve rice varieties to different biotypes of BPH. It was shown that light intensity, seedling age, and nitrogen application rate had significant effects on the resistance performance and stability of rice varieties to different BPH biotypes. Among the varieties with resistance to BPH biotype II, the RHT, RP1976-18-6-4-2, and Ptb33 showed stable, IR56 showed unstable, while IR36 and ASD7 showed the most unstable resistance; whereas among the varieties susceptible to BPH biotype II, the TN1 showed stable, Guihuazhan, Foshanyouzhan and IR26 showed relatively stable, while Guojing No. 4 and Mudgo showed unstable susceptibility. For the rice varieties resistant to BPH biotype Bangladesh, the RHT, RP1976-18-64-2 and Ptb33 presented unstable, and IR56 presented very unstable resistance; whereas for the varieties susceptible to BPH biotype Bangladesh, the Guihuazhan, Foshanyouzhan and IR26 performed stable, TN1 and IR36 performed unstable, and Guojing No. 4, Mudgo and ASD7 performed the most unstable susceptibility.