| Summary: | Rice is susceptible to cold stress at the seedling stage, which can delay growth and decrease yield. We evaluated 187 rice accessions for cold tolerance at the seedling stage and developed genotypic assays for three markers. All <i>japonica</i> (20/20) and 20/140 <i>indica</i> accessions were highly cold tolerant. Two SNP markers specific for <i>COLD1</i> and <i>LOC_Os10g34840</i> were practical to use by normal agarose gel. The SNP marker specific for <i>COLD1</i> was highly specific for predicting cold tolerance. However, the sensitivity of this marker was low as several cold-tolerant <i>indica</i> accessions lacked the cold-tolerant allele. The <i>LOC_Os10g34840</i> marker was slightly more sensitive than the <i>COLD1</i> marker for predicting highly cold-tolerant accessions. An insertion/deletion variant in the <i>NAC6</i> gene was identified as a novel cold tolerance marker. The <i>NAC6</i> marker predicted more highly cold-tolerant accessions compared with the other two markers. The SNP marker specific for <i>LOC_Os10g34840</i> and the <i>NAC6</i> marker were present in several tested subgroups, suggesting their wide effects and distribution. The three markers combined predicted the most highly cold-tolerant accessions, indicating that the marker combination is superior for applications such as marker-assisted breeding. The cold-tolerant accessions and the genotypic marker assays will be useful for future rice breeding.
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