Physiological Characteristics And Transcriptome Analysis Of Apple Rootstock Seedlings Malus Halliana Under Iron Deficiency Stress

Iron?Fe?is an essential nutrient for plants.Fe deficiency will lead to blocked synthesis of chlorophyll synthetic precursors,the pyrrolidine ring and porphyrin ring,the photosynthesis rate decreasing,chlorosis in leaves,even whole plant decay of fruit trees.Utilization of Fe deficiency-tolerant rootstock is an effective strategy to prevent Fe deficiency problems in fruit trees production.Malus halliana grows well and the chlorotic symptom associated with Fe deficiency in apple trees was not found on the cultivar in the Northwest Loess Plateau of China.M.halliana showed characteristics of Fe deficiency-tolerant rootstocks.However,few studies on resistance have been conducted on M.halliana.In this experiment,the Fe deficiency stress was carried out under the hydroponic condition,and the mechanism of Fe deficiency stress tolerance was preliminarily revealed from the physiological level and the transcriptional level.Main results are as follows:1.Under Fe deficiency,the contents of chlorophyll a,chlorophyll b and carotenoid in the leaves of M.halliana increased firstly and then decreased,the relative conductivity increased gradually,the content of proline increased significantly in 0.5d,the activity of POD enzyme increased gradually,the activity of APX enzyme increased significantly,the pH value of rhizosphere decreased and the root activity gradually increased.Based on the above results,0d?T1?,0.5d?T2?and 3d?T3?were selected as transcriptional sequencing points.2.In all,6907,5328,and 3593 differentially expressed genes?DEGs?were identified in pairs of T2 vs.T1,T3 vs.T1,and T3 vs.T2.Several of the enriched DEGs were related to heme binding,Fe ion binding,thylakoid membranes,photosynthesis-antenna protein,porphyrin and chlorophyll metabolism and carotenoid biosynthesis under Fe deficiency.Fe deficiency induced significant down-regulation in many genes involved in photosynthesis at T2 and a strong up-regulation of most photosynthesis genes was detected at T3.A similar expression pattern was found in pigment regulation.M.halliana can restore photosynthesis under Fe deficiency.3.In roots,2473,661,and 776 DEGs were identified in pairs of T2 vs.T1,T3 vs.T1,and T3 vs.T2.We found that several DEGs were enriched in photosynthesis,glycolysis and gluconeogenesis,tyrosine metabolism and fatty acid degradation pathways.Under Fe deficiency,the expression levels of genes involved in glycolysis pathwaywereup-regulatedincluding fructose-1,6-bisphosphatase?FBPase?,6-phosphofructokinase?PFK?,phosphoglycerate kinase?PGK?and pyruvate kinase?PK?.Fe deficiency significantly affected sugar metabolism in roots of M.halliana.4.To ensure the accuracy of transcriptome data,we verified sixteen genes related to photosynthesis-antenna protein,porphyrin and chlorophyll metabolism and carotenoid biosynthesis pathways in leaves and fifteen genes involved in glycolysis and sugar synthesis,metabolism and transportation in roots using quantitative real-time PCR?qRT-PCR?.The relative expression of 28 genes was significantly correlated with transcriptome sequencing results.5.In view of the KEGG analysis that the DEGs in leaves were mainly enriched in photosynthesis pathway,chlorophyll fluorescence parameters,photosynthetic parameters and chlorophyllase?Chlase?activity were measured.Under Fe deficiency,the minimum fluorescence?F0?,maximum fluorescence?Fm?,effective quantum yields?Y?II??and quantum yield of regulated energy dissipation?Y?NPQ??increased and electron transport rates?ETR?decreased.Under Fe deficiency,Net photosynthetic rate?Pn?decreased,and stomatal conductance?Gs?,transpiration rate?E?and intracellular CO₂ concentration?Ci?increased.The activity of chlase decreased in T2and increased in T3.6.The contents of glucose,fructose and sucrose in roots and leaves were measured.Under Fe deficiency,sucrose was accumulated in roots and leaves.The content of glucose in roots decreased significantly and had no significant change in leaves.The content of fructose decreased significantly in leaves and no significant change in roots.Sugar synthesis,utilization and distribution are affected by Fe deficiency.Sugar regulation is a way of M.halliana to respond to Fe deficiency stress.