Study On Single Nucleotide Polymorphisms (SNPs) Associated With High Use Efficiency Of Water And Light Resources In Populus Nigra L.

Populus nigra L. is an important gene donor for major poplar varieties worldwide, and plays crucial roles in the genetic improvement programs of poplar in China. There is a serious shortage of gene resources of Aigeiros section in China, as P. nigra distributes only across a small area of Northern Xinjiang. In recent years, many P. nigra genotypes were introduced from 15 European countries to China successively, making gene resources of P. nigra substantially increased in our country. China is a nation with a shortage of fresh water, resulting in restrained growth and production of poplar plantations due to insufficient water supply for land soils. Thus, breeding water economic and drought resistance poplar variety by improving its water use efficiency (WUE) and the capacity of drought tolerance is important for poplar production and conservation of environments. Photosynthesis is the key factor that impact biomass accumulation of plants. This makes the breeding of poplar variety with higher photosynthetic-light use efficiency (PUE) is a key point in the establishment of high productive plantations. Such varieties should have high ability of the utility of light resource.In this paper, gene resources of P. nigra collected from Europe and Xinjiang were used as materials. PUE was evaluated and single nucleotide polymorphisms (SNPs) within water and photosynthesis-related genes were discovered and analyzed combined with the results of evaluation of WUE (δ¹³C) studied previously. Functional SNP markers were found by using association analyses between SNPs and related traits, proving a foundation for marker-assistant breeding on water-use and photosynthesis traits in poplar. The main results are described as following:Evaluation of WUE and PUE in P. nigra. Since the evaluation of WUE has been done in previous work, this paper focused on the evaluation of PUE. Gas exchange and chlorophyll fluorescence-related parameters were measured in gene resources of P. nigra. The results showed abundant genetic variation for traits of photosynthesis and growth in P. nigra. The genotypes from China and the U.K. have relatively higher values of photosynthesis rate (Pn), while genotypes from Hungry have the highest values of quantum efficiency of chloroplast photosystem II (PSII) centers (Fv/Fm). The measured traits were found to have generally high broad sense heritabilities (H2 ranges from 0.750 to 0.962) with considerable variation coefficients. The results showed that parameters including Pn, intercellular concentration of carbon dioxide (CO2) (Ci), stomatal conductance (Gs) and electron transport rate (ETR) were significantly correlated with height growth, indicating potential applications for the evaluation of PUE in P. nigra. Focus should be on genotypes from counties of Southern and Eastern Europe in further breeding research based on PUE in poplar.Discovery and analysis of SNPs from genes that are involved in water and photosynthesis use. A total of 312 single nucleotide polymorphism (SNP) sites are found from seven water-use related (SuSY1, dhn, lea3, PIP, Expa1, ERF, and GPX1) and two photosynthetic-light use related (rubisco and Lhcb2) genes. The median SNP frequency is one site per 26 bp. The average nucleotide diversity for the sequenced regions was calculated to beθ_W=0.01074 andÏ€_T=0.00702, higher values than those observed in P. tremula, P. trichocarpa and most conifer species. Tests of neutrality for each gene reveal a general excess of low-frequency mutations, and many non-synonymous polymorphisms of these loci might be involved in local adaptation in P. nigra. The results reject neutral evolution at GPX1, dhn, lea3, and Expa1. These loci are likely to be undergoing a selective sweep which can cause hitchhiking effect. The level of linkage disequilibrium (LD) varies among loci. The overall LD in P. nigra is low, decaying rapidly from 0.45 to 0.20 or less within a distance of 300 bp. This is similar to the rate of decay reported in most other tree species, indicating that P. nigra is suitable for association analysis and mining of functional SNP markers based on candidate genes.Association analysis between SNPs and WUE trait (δ¹³C value). A total of 25 SNPs were genotyped using SNaPshot^? technology. These SNPs are from five genes, namely SuSY1, lea3, PIP, Expa1, and ERF. Association analysis between marker and trait were carried out using both One-way Analysis of Variance (ANOVA) and General Linear Model (GLM). The results showed six associations between markers andδ¹³C value. SNP2 shows T-C transition with a genetic effect of 7.705%. The trees with genotype TT at SNP2 have higherδ¹³C values than that for other genotypes. SNP15 shows G-A transversion with a genetic effect of 7.570%. SNP15 causes an amino acid alteration between Val and Ile, and genotype GG has higherδ¹³C value. SNP25, SNP26, SNP27 and SNP31 are all located in Expa1, with a genetic effect ranging from 6.436% to 6.900%. One genotype was not detected for each of the four SNPs, and similar patterns of genotype frequency was found among them. For SNP25, SNP27 and SNP31, genotype TT has higherδ¹³C value than heterozygous genotype, implying similar mechanism responsible for the variation ofδ¹³C value in P. nigra.Association analysis between SNPs and PUE related traits. A total of 20 SNPs were genotyped using SNaPshot? technology. These SNPs are from genes of rubisco and Lhcb2. Association analysis between marker and traits were carried out using both ANOVA and GLM. The results showed that 4 SNPs are closely associated with photosynthesis and growth traits. SNP12, SNP36 and SNP50 are all located in rubisco. Among them, SNP12 and SNP36 are non-synonymous mutations, which result in amino acid alterations of Asn-His and Tyr-Phe, respectively. SNP12 and SNP36 are associated with ETR, and the genotype AA has higher ETR value than other genotypes. SNP50 is associated with Pn, ETR, Fv/Fm, height and diameter, and genotype CT has higher values of ETR, Fv/Fm, height and diameter and lower value of Pn. SNP44 of Lhcb2 is associated with height growth, and the genotype GG has higher height growth than other genotypes.