Identification of stay-green candidate gene TaTRNH1-3B and development of molecular markers related to chlorophyll content and yield in wheat (Triticum aestivum L.).

Functional stay-green characteristic is closely associated with delayed loss in photosynthetic function and increased crop yield. However, the development and application of functional molecular markers based on stay-green-related genes are limited. This study compared and analyzed the differences of SPAD values, photosynthetic parameters, fluorescence parameters, and antioxidant enzyme activities at 0, 10, 18, 22, 26, 30 and 34 days after anthesis, as well as agronomic traits at mature stage between a stay-green line, Tailv113 (TL113), and a non-stay-green cultivar, Jinmai39 (JM39). The results showed that TL113 had higher photosynthetic capacity, photosynthetic efficiency, antioxidant capacity and yield than JM39. Subsequently, a comparative transcriptome analysis was conducted on TL113 and JM39 at 0, 26, and 30 days after anthesis. Analysis showed that senescence-associated co-expressed genes (SCEGs) and stay-green-associated differentially expressed genes (SDEGs) jointly affected wheat leaf senescence, while SDEGs played an important role in the stay green differences between TL113 and JM39. By analyzing the SNP sites of the SDEGs from transcriptome sequencing, a nsSNP was found in the TaTRNH1-3B sequence between TL113 and JM39. Further analyzing the resequencing data published in the Wheat Union database, four linked SNP sites were identified in TaTRNH1-3B, which formed two haplotypes, TaTRNH1-3B-Hap1 and TaTRNH1-3B-Hap2. Based on the SNP at 373 bp (A/G), a CAPS molecular marker, TaTRNH1-3B-Nla III-CAPS, was developed to distinguish allelic variations (A/G). Association analysis between TaTRNH1-3B allelic variation and agronomic traits found that the accessions possessing TaTRNH1-3B-Hap1 (A) exhibited significantly higher SPAD values than those possessing TaTRNH1-3B-Hap2 (G) in 6 of 10 environments at the jointing stage and in 7 of 10 environments at the grain filling stage in Beijing. Similarly, the accessions possessing TaTRNH1-3B-Hap1 (A) exhibited significantly higher chlorophyll contents and yield than those possessing TaTRNH1-3B-Hap2 (G) in 3 environments in Taigu. Additionally, lines with TaTRNH1-3B-Hap1 (A) displayed higher SPAD values at 0, 15, and 20 days after anthesis in the two BC3F3 populations than those with TaTRNH1-3B-Hap2 (G). These results suggest that TaTRNH1-3B is associated with the stay-green and yield traits in wheat, and TaTRNH1-3B-Hap1 is a favorable stay-green haplotype. The newly developed molecular marker, TaTRNH1-3B-Nla III-CAPS, provide valuable information for wheat genetic improvement of stay-green and high-yield traits, and can be used to marker-assisted selection breeding.