**5. Conclusions**

In the present study, we performed QTL analysis and identified QTLs for LTG and coleoptile length in the F2 population. Among the five QTLs for LTG, two major QTLs, *qLTG1* and *qLTG3*, and three minor QTLs were detected on chromosomes 8 and 10. Fine mapping revealed that two QTLs, *qLTG10.1* and *qLTG10.2*, were linked on chromosome 10 and exerted opposite effects with the Hwaseong allele at *qLTG10.2* and the *O. rufipogon* at *qLTG10.1*, respectively, in turn, increasing LTG. Because two LTG QTLs, *qLTG10.1* and *qLTG10.2*, which act in opposite directions are linked, the DNA markers could improve the selection efficiency of the high LTG lines with *qLTG10.1* from *O. rufipogon* and *qLTG10.2* from Hwaseong. Interactions among QTLs were not significant, implying that the QTLs act in an additive manner. NIL plants with combinations of favorable alleles from *O. rufipogon* and Hwaseong exhibited the highest LTG among all groups, supporting the absence of interactions. With regard to coleoptile length, three QTLs observed on chromosomes 1, 3, and 8 were colocalized with QTLs for LTG, suggesting the pleiotropy of the single gene at each locus. According to the results, the introgression of favorable *O. rufipogon* alleles should hasten the breeding of high LTG and coleoptile elongation in *japonica* cultivars.

**Supplementary Materials:** The following are available online at http://www.mdpi.com/2073-4425/11/10/1200/s1, Figure S1: Measurement of low-temperature germination rate of parental lines (*O. rufipogon*, Hwaseong, CR1517 and CR1518) for 4–8 days after incubation (DAI). Seeds were considered as germinated when coleoptile emerged. Arrows indicate emerged coleoptile, Figure S2: Comparison of germination rates of *O. rufipogon* (Ru), Hwaseong (Hs), CR1517 and CR1518 at normal (30 ◦C) (A) and low-temperature conditions (13 ◦C) (B). Different letters at each DAI indicate significant difference at *p* < 0.05 based on Tukey's test. ns: no significant. Error bars indicate standard deviation, Figure S3: Interaction analysis of *qLTG1* and *qLTG3* with other QTL for LTG based on LTG scores of the F2 population. HH, HR and RR indicate Hwaseong homozygous, heterozygous (Hwaseong/*O. rufipogon*) and *O. rufipogon* homozygous at two QTLs, respectively. Low-temperature germination rates at 7 days after incubation were compared. Error bars indicate standard deviations. Interaction was determined by general linear regression model, Figure S4: Comparison of coleoptile length of parental lines at normal (30 ◦C) (A) and low-temperature (13 ◦C) (B) Error bars at each DAI indicate standard deviation. Different letters at each DAI indicate significant difference at *p* < 0.05 based on Tukey's test. ns: no significant. Ru: *O. rufipogon*; Hs: Hwaseong, Table S1: List of primers used in this study, Table S2: Comparison of germination rates among parental lines and pyramiding lines at 6 and 7 DAI.

**Author Contributions:** M.A. performed experiments and analyzed data. M.A., K.-C.S., C.A., and S.-N.A. conceived the study, designed, and supervised the study. H.-S.L., S.H.K., Y.-AJ., C.A., N.H.L., J.-Y.L. and J.-W.K. investigated the traits. M.A., and K.-C.S. wrote and C.A. and S.-N.A. edited the manuscript. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research was carried out with the support of "Cooperative Research Program for Agriculture Science and Technology Development (Project No. PJ01321401)", Rural Development Administration and of Korea Institute of Planning and Evaluation for Technology in Food, Agriculture, Forestry and Fisheries (IPET) through Golden Seed Project funded by the Ministry of Agriculture, Food and Rural Affairs (MAFRA) (213009-05-3-WT312), Republic of Korea.

**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.
