| 摘要: |
| 为初步解析马尾松花粉对超低温保存的生理响应及相关代谢机制，本项目以马尾松花粉为研究对象，在-196℃液氮进行超低温保存并分析冷存过程中（冻存前CK，液氮冻存后LN，化冻后HD）与ROS相关的生理指标及转录组数据。结果表明：（1）马尾松花粉超低温保存的最适含水量为3.96%，该含水量下将花粉直接投入液氮保存48h，冻存前后花粉存活率分别为78.54%和73.80%；（2）SOD，GSH，APX，抑制羟自由基能力4个指标在冻存前后差异显著；（3）转录组测序共获得65.60Gb过滤数据，有38，505个基因比对到参考基因组（47.84%），CK&LD，CK&HD，LD&HD的差异表达基因（DEGs数量分别为232、268和218个。GO和KEGG分析表明，应激响应（Response to stimulus）、抗氧化活性（Antioxidant activity）等GO term显著富集；植物激素信号转导、MAPK信号途径、淀粉和蔗糖代谢、果糖和甘露醇代谢、过氧化物酶体等KEGG途径显著富集，从这些途径中进一步筛选到10个可能与冷冻保存过程中损伤与修复有关的基因。研究结果为马尾松种质资源高效利用和安全保存提供了技术手段，也为进一步解析冷冻保存损伤的分子机制提供参考。 |
| 关键词: 马尾松，花粉，超低温保存，生理，转录组 |
| DOI：10.11931/guihaia.gxzw202403015 |
| 分类号: |
| Fund project:广西自然科学基金（2021GXNSFAA196069&2016GXNSFBA380224）；广西优良用材林资源培育重点实验室项目（2020-A-03-01）；广西重点研发计划项目（桂科AB24010291） |
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| Transcriptome analysis and physiological response to cryopreservation of Pinus massoniana pollen |
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ZHANG Xiaoning1, ZHANG Ye1, XU Zhanwu2, QIN Zihai1, WEI Qiulan1, LIN Dong1, XIAO Yufei1, LIU Hailong1*
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1.Guangxi Key Laboratory of Superior Timber Trees Resource Cultivation, Guangxi Forestry Research Institute, 530002, Nanning , China；2. State Key Laboratory of Biocatalysis and Enzyme Engineering, College of Life Sciences, Hubei University, 430062, Wuhan, China
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| Abstract: |
| This study aims to investigate the response mechanism of Pinus masson pollen to ultra-low temperature, to enhance its preservation and utilization efficiency. Masson pine pollen was cryopreserved and the physiological indexes related to reactive oxygen species (ROS) were assessed at different stages (ck before freezing, LN after freezing, HD after freezing and de-freezing). Transcriptome sequencing was also performed using the Illumina HiSeq TM 6000 platform. Results indicated that: (1) The optimal water content for cryopreservation was 3.96%, with survival rates of 78.54% before freezing and 73.80% after post-freezing. (2) During the frozen storage process significant differences were observed in the four indexes of Superoxide Dismutase (SOD), Glutathione (GSH), Ascorbate Peroxidase (APX), and the ability to inhibit hydroxyl free radicals, which are related to oxidative stress induced by ROS. (3) A total of 65.60Gb of Clean Data were obtained, with 38,505 genes mapped to the reference genome (47.84%); The number of differentially expressed genes (DEGs) in comparisons between control and freezing treatment (CK&LD), control and de-freezing treatment (CK&HD), and freezing and de-freezing treatment (LD&HD) were 232, 268, and 218, respectively. Additionally, Gene Ontology (GO) and Kyoto Encyclopedia of Genes (KEGG) annotation revealed significant enrichment of GO term such as response to stimulus and Antioxidant activity, and KEGG Pathway such as Plant hormone signal transduction, MAPK signaling pathway, Starch and sucrose metabolism, Fructose and mannose metabolism, and Peroxisome pathways. 10 genes screened in these pathways may be closely related to freezing and de-freezing process. The results help to improve the efficient of preservation and utilization of the Masson Pine pollen, and also provided reference for further analysis of the molecular mechanism of its freezing mechanism. |
| Key words: Pinus massoniana, pollen, crypreservation, physiologica, Transcriptome sequencing. |
