Abstract: [Objective] The characteristics of pollen germination and the differences of allergenic proteins among six poplar germplasms (Populus deltoides ‘2KEN8’, P. deltoides ‘Nanyang’, P. deltoides×P. maximowiczii ‘Eridano’, P. deltoides×P. cathayana ‘Senhai 2’, P. simonii and P. nigra var. italica) were studied to provide a theoretical basis and scientific evidence for breeding hypoallergenic poplar varieties and for the control of plant allergy.[Method] Pollen was collected from flower branches of six poplar germplasms collected from Beijing, Hebei and Henan in March 2015 for a period of hydroponic culture, pollen germination was studied by liquid culture method, and pollen allergenic proteins of different poplar germplasms was used by two-dimensional electrophoresis and real-time quantitative PCR.[Result] 1) The pollen viability of P. deltoides ‘2KEN8’, P. deltoides ‘Nanyang’ and P. nigra var. italica was higher, followed by P. simonii and P. deltoides×P. maximowiczii ‘Eridano’, however, the viability of P. deltoides×P. cathayana ‘Senhai 2’ was the worst; the largest pollen germination rate was in P. deltoides ‘2KEN8’, up to 74.42% (±2.36%), while the pollen germination of triploid P. deltoides×P. cathayana ‘Senhai 2’ was 0; the average length of pollen tube of P. nigra var. italica was 456 μm (±2.05 μm), the shortest was in P. simonii, 276.44 μm (±11.08 μm), and the difference of pollen germination rate and pollen tube length among different entities were highly significant (P<0.01), suggesting that the pollen viability was significantly different among different germplasms. 2) The 2-DE maps of P. deltoides ‘Nanyang’, P. deltoides×P. maximowiczii ‘Eridano’, P. deltoides×P. cathayana ‘Senhai 2’, P. simonii and P. nigra var. italica mature pollen were analyzed by Imagemaster 2D platinum 6.0, a total of 408 protein spots were matched. Based on the 28 potentially allergenic proteins of P. deltoides ‘2KEN8’ identified in our previous study, 6 allergenic proteins were missing expression in the other five entities; both P. nigra var. italica and P. deltoides ‘Nanyang’ had 21 allergenic proteins, P. deltoides×P. maximowiczii ‘Eridano’ had 19; while P. deltoides×P. cathayana ‘Senhai 2’ and P. deltoides ‘2KEN8’ had the least number of allergenic proteins, only 18. Morover, the analysis of difference expression indicated that 12 of the 22 remaining allergenic proteins were less expressed in the five entities than in P. deltoides ‘2KEN8’, the range of relative expression level was -3.99——0.02. The expression level of P. deltoides×P. cathayana ‘Senhai 2’ and P. simonii was the least, especially they had 10 and 8 allergenic proteins which were significantly or highly significant lower than that of P. deltoides ‘2KEN8’, respectively. The expression of 3 allergenic proteins in the 5 entities was higher than in P. deltoides ‘2KEN8’. The strongest expression was in P. deltoides ‘Nanyang’, significantly greater than in P. deltoides ‘2KEN8’, followed by P. nigra var. italica, P. deltoides×P. cathayana ‘Senhai 2’, P. deltoides×P. maximowiczii ‘Eridano’ and P. simonii. Variance analysis among groups suggested that the expression of 17 allergenic proteins of P. nigra var. italica and P. deltoides ‘Nanyang’ were significantly (P<0.05) or highly significantly (P<0.01) greater than that of P. deltoides×P. cathayana ‘Senhai 2’ and P. simonii. 3) qRT-PCR results showed that there were 5 of 9 allergenic proteins encoding genes highly expressed in P. nigra var. italica and P. deltoides ‘Nanyang’, including spot 12, 51, 200, 216 and 161, followed by P. simonii, while both of P. deltoides×P. maximowiczii ‘Eridano’ and P. deltoides×P. cathayana ‘Senhai 2’ had only 2 highly expressed allergenic protein coding genes.[Conclusion] The pollen viability, the number and expression levels of allergenic proteins were significantly different among the 6 germplasm entities, providing an important basis for future breeding of new poplar hypoallergenic varieties. However, further study is needed to understand the correlation between expression levels and allergenicity of the allergenic proteins of mature pollen.

Key words: Populus, mature pollen, allergenic protein, two-dimensional electrophoresis, real-time quantitative PCR