Genome-wide Association Study Of K Content And The Other Correlated Traits Under Different K Application Rates

Potassium is one of inorganic nutrients required by plants for growth and development,playing a crucial role in growth,metabolism,enzymatic activity and osmotic adjustment of plants.The available nutrients are deficit seriously in soils in China at present,and the potash is deficiency in agricultural soils widely.The low potassium efficiency in tobacco-growing soils makes it difficult to significantly raise the potassium content in tobacco leaves.The potassium content in tobacco leaves closely related to tobacco quality is an important indicator of tobacco quality evaluation.Studies have shown that the absorptive capacity of potassium and potassium content in leaves differ among genotypes.In case of the soil potassium deficiency,it is a practical and feasible way to promote high quality tobacco leaves production in China by breeding of new tobacco varieties with high potassium content through tapping into crop potential to increase efficiencies of potassium nutrient.In this research,the 92 tobacco germplsm resources were chosen to investigate the phenotypic differences of K content and other relevant traits in tobacco leaves.Field experiments were carried out for 2 years?2014-2015?underK300?K2O=300 kg/hm2?and K150?K2O=150 kg/hm2?,in the main tobacco producing region,Xichang city,Liangshan Prefecture,Sichuan Province.Meanwhile the genetic diversity and structure of tested tobacco population was analyzed with SNP.Then we used a large amounts of SNPs to combine existing phenotype data into genome-wide association study?GWAS?based on the previous genetic analyses trying to decipher the genetic architecture attributed to potassium content and related traits in tobacco leaves.The research contents and results were listed as follows:?1?The potassium content in leaves were decided collaboratively by potash application and genotypes,the results showed that there were significant differences in K content among the genetypes under the two K levels.The range of K content was 1.30%-3.62%and 0.19%-2.48%?CV2014=20.34%,CV2015=36.16%?in upper leaf,1.37%-3.73%and 0.69%-3.30%?CV2014=17.31%,CV2015=28.66%?in middle leaf,2.13%-4.65%and 0.75%-6.14%?CV2014=14.63%,CV2015=33.78%?in lower leaf under K300.While under K150,the range of K content in upper,middle and lower leaf were respectively 0.88%-2.80%and 0.31%-1.77%CV2014=20.18%,CV2015=27.18%),1.24%-2.88%and 0.37%-3.09%?CV2014=15.81%,CV2015=32.92%?,1.69%-4.95%and 0.44%-3.68%?CV2014=20.34%,CV2015=27.70%?.The K content under normal K level was significantly higher than that under low K level.The average K content in different leaf positions under normal K rate were respectively 1.13,1.14,1.15?2014?and 1.10,1.25,1.35?2015?times that under less K rate.The K content was higher under normal K rate,indicating that K application affected on the K content in leaves,but the effect was lower than genotypes itself.?2?There were significant differences of the chemical compositions among genetypes as well under the two K application rates.In the two-years field experiment,the content of the total sugar,nicotine,reducing sugar,chlorine and the total nitrogen in tobacco leaves of different stalk positions varied from 5.11%?29.31%,0.80%?3.48%,2.59%?14.71%,0.15%?0.49%,1.23%?3.26%respectively under K300,and varied from 8.35%?29.59%,0.84%?3.61%,7.96%?13.97%,0.15%?0.28%,1.49%?2.43%respectively under K150.The highest values of the five chemical components 5.74,4.35,5.68,3.27,2.65 times and 3.54,4.30,1.76,1.87,1.63 times the lowest under K300 and K150.?3?93685 SNPs spreading across the tobacco genome were developed based on the simplified development of genome sequencing,and then the SNPs were applied to the follow-up study.The average value of genetic diversity index of the tested germplasm was 0.41,stating that the population had a relative low genetic diversity.Genetic structure analysis showed that the association panel could be divided into 4 subgroups and gene exchange occurred between those subgroups.To minimize the effect of environmental variation,phenotypic BLUP?best linear unbiased prediction?values across two K rates and two years were used for association studies.Then we used the 93685 SNPs over the whole genome with the breeding values of potassium content into genome-wide association analysis.The results showed that there were 54,621 and 663 SNPs the numbers of candidate genes were 45?490?527 respectively that associated with K content in upper,middle and lower leaves.And there were 3 SNPs associated with K content in upper,middle and lower leaves simultaneously.The 6 candidate genes associated with these 3 SNPs may be related to the development and stress resistance of plant.?4?There were 2130 reliable SNPs and 1433 candidate genes associated to the breeding values of chlorine,nicotine,reducing sugar and total sugar through genome-wide association study at both K rates,and the annotations of candidate genes were displayed.There was only one candidate gene associated to the chlorine,reducing sugar and total sugar simultaneously,and the numbers of candidate genes associated to total sugar with reducing sugar,total nitrogen,and chlorine respectively were 1,418 and 1.One SNP marker named chrNW₀15945306.1₂8081 associated with chlorine content in middle and lower leaves simultaneously can well explain the phenotypic variation with a high R2?R2=48.90?68.92?.Another SNP chrNW₀15929377.1₁19905 associated with reducing sugar?R2=0.01?and total sugar?R2=0.57?simultaneously,indicating that there may be genetic linkage between the two traits or pleiotropism.Through the interpretation of the candidate gene associated with quality chemical characters to find the similar or homologous genes,we found that found that most of these candidate genes involved in the metabolic network mechanism including plant growth and development,signal transduction,cold resistance and drought adversity stress resistance.