*4.7. qRT-PCR and Expression Analysis*

The extraction of the total RNA was carried out with a TRIzol reagent. The first-strand of cDNA was reverse transcribed by using a TransScript II First-Strand cDNA Synthesis SuperMix kit (Transgen). The qRT–PCR analysis was executed by using the kit named SYBR FAST qPCR (KAPA). The

#### qPCR primer pairs sequences for *LOC\_Os06g01320* were 5- -CAAAAAAAAAGACAATAAGGTGGA-3- (forward) and 5- -CAGACATTGCTTACCCTTATTTATTTT-3- (reverse). EF-1 alpha was used as an internal control, and the sequencing was 5- -GCACGCTCTTCTTGCTTTCACTCT-3- (forward) and 5- -AAAGGTCACCACCATACCAGGCTT-3-(reverse).

**Author Contributions:** S.J., L.W., X.Y., J.W. conducted the phenotyping and genotyping of parental lines and individuals of the RIL population. C.Y. and S.J. performed the SNP calling, data analyses, and bin-map construction. X.Z., X.S., Q.X., B.L., H.L., and W.L. participated in the phenotyping and data analysis. S.J., Z.L., and W.L. designed the experiment. S.J., C.Y., Q.X. and X.S. drafted the manuscript. All authors have read and agreed to the published version of the manuscript.

**Funding:** This work was supported by a grant from the National Key Research and Development Program of China (2016YFD0300104), the National Natural Science Foundation of China (31661143012), the Natural Science Foundation of Heilongjiang Province of China (C2016050, YQ2019C020), Heilongjiang Postdoctoral Financial Assistance (LBH-Q15133), Provincial funding for the National Key Research and Development Program in Heilongjiang Province (768001), and Heilongjiang Province Agricultural Science and Technology Innovation Project (2018CQJC002, 2019CQJC002).

**Conflicts of Interest:** The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.
