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Drought stress is one of the major agronomic concerns that lead towards a sharp decline in sugarcane yield. An urgent demand to overcome drought is critical to ensure sugarcane production. Mutation breeding is one of the promising tools available to produce stress-resistant plants, with the induction of new alleles due to point mutation within existing sugarcane germplasm. The current study was directed to chemically mutagenize the calli of two sugarcane cultivars (ROC22 and FN39) via 0.1% EMS, with focus on inducing mutations in their genome. The 1644 regenerated plants of ROC22 and 1398 of FN39 were exposed to 28% PEG-6000 stimulated osmotic stress. Eighteen plants of ROC22 and 2 plants of FN39, that survived after in vitro osmotic stress treatment, were then subjected to preliminary greenhouse pot trials to confirm drought tolerance by analyzing them using various physiological parameters, including photosystem II (PSII) photochemical efficiency (Fv/Fm), leaf chlorophyll content, and photosynthetic rate. The genetic diversity among drought-resistant mutant lines was further assessed by 15 pairs of simple sequence repeat (SSR) markers amplification and CEL (Celery) I endonuclease digestion, to investigate the mutated sites. Mutant lines of ROC22 (i.e., MR22-15 and MR22-20) were found to be promising for future drought resistance breeding, due to better physiological adaptation under drought stress.

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Background: The NAC acronym originates from these three genes (NAM, ATA1/2 and CUC2), and were proved to participate in plant development, sugar accumulation and stress tolerance. However, little information is available regarding these genes in sugarcane. The newly published sugarcane genome provided an opportunity to identify the SsNAC transcription factors. Results: In this study, a total of 151 NAC genes including 327 alleles were identified in the autopolyploid S. spontaneum genome and were distributed unevenly on eight chromosomes with the majority located on Chr 5(54, 16.51%), Chr 1(51, 15.60%) and Chr 2(51, 15.60%). 71.6%(234) and 12.53%(41) of SsNAC genes were mainly derived from segmental duplication and tandem duplication. Phylogeny analysis of NAC TF proteins by comparing NAC between sugarcane and Arabidopsis thaliana suggested that the NAC family can be divided into 15 subgroups with one subgroup unclassified. RNA-seq data analysis revealed that 82 SsNAC were expressed in 15 segments of the developmental gradient of the leaf and 74 SsNAC were presented in 12 different developmental stages, respectively. Remarkably, SsNAC genes of the ATAF subgroup presented the highest expression levels among the subgroups, and seven of eight SsNAC genes from the ATAF subgroup excluding SsNAC30 were present at much higher expression levels in the developmental gradient of the leaf than those in different developmental tissues types, indicating that the ATAF subgroup may have significant roles and participate in leaf growth and development and photosynthesis. Importantly, the NAC1 subgroup SsNAC91 gene, being orthologous to Sobic.006G147400 ( Dry gene ), displayed significantly higher expression levels in premature and mature stems of the low sucrose and low water content species S. spontaneum as opposed to the orthologous gene in the high sugar and high water content species S. officinarum . This suggests that the SsNAC91 gene may also play important roles in cellulose biosynthesis and water transport in sugarcane as it does in sorghum. Conclusions: This study provided the basis for the comprehensive genomic study of the SsNAC gene family and thus established a good foundation for the functional analyses of SsNAC genes which can be utilized to breed new varieties of sugarcane.

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