Germplasm Innovation and Characteristic Analysis of Transgenic PsnNAC007Populus simonii×P. nigra with High Drought Tolerance

doi:10.7525/j.issn.1673-5102.2024.03.004

Abstract

Abstract:

Populus simonii×P. nigra is a fast-growing and cold-tolerant tree species with elite wood properties in northeast China. In order to produce a new forest germplasm with more widely planting and improved drought tolerance， this study successfully generated transgenic P. simonii×P. nigra using PsnNAC007， a key drought response transcription factor gene. Growth characteristics， drought tolerant ability， net photosynthetic rate（P_n）， transpiration rate（T_r）， stomatal conductance（G_s）， water conductance， cell morphology and wood components of overexpressed PsnNAC007 transgenic plants were analyzed respectively. The results showed that there was no significant difference in growth between transgenic and wild-type plants， but drought survival rate was increased by 26.15%. Under drought condition， transgenic plants exhibited reduced Cleaf and T_r， improved water use efficiency， and lower water conductance loss. Anatomical analysis showed that the overexpression of PsnNAC007 led to more and smaller stem xylem vessels， which were conducive to the continuous and efficient water transport in plants under drought conditions. Based on wood components analysis， lignin deposition in stems of transgenic plants was increased significantly， and monosaccharides that constituted cellulose and hemicellulose showed no significant change.

Key words: Populus simonii × P. nigra, drought stress, transcription factor, genetic transformation

CLC Number:

S792.11

Xiaoqian WU, Xu HE, Jinghui GAO, Shuang LI. Germplasm Innovation and Characteristic Analysis of Transgenic PsnNAC007Populus simonii×P. nigra with High Drought Tolerance[J]. Bulletin of Botanical Research, 2024, 44(3): 349-360.

Figures/Tables 12

Table 1

Fig.1

Fig.2

Fig.3

Fig.4

Fig.5

Fig.6

Fig.7

Fig.8

Fig.9

Fig.10

Table 2

References 33

1	山仑，邓西平，康绍忠.我国半干旱地区农业用水现状及发展方向［J］.水利学报，2022（9）：27-31.
	SHAN L， DENG X P， KANG S Z.Current situation and perspective of agricultural water used in semiarid area of China［J］.Journal of Hydraulic Engineering，2022（9）：27-31.
2	许恩银，聂影，芮晓东.基于森林资源清查数据的林地利用效率变化研究［J］.南京林业大学学报（自然科学版），2022，46（5）：213-220.
	XU E Y， NIE Y， RUI X D.Analysis on the forest land use efficiency changes based on forest resource inventory data［J］.Journal of Nanjing Forestry University （Natural Sciences Edition），2022，46（5）：213-220.
3	BANDURSKA H.Drought stress responses：coping strategy and resistance［J］.Plants，2022，11（7）：922.
4	BRÉDA N，HUC R， GRANIER A，et al.Temperate forest trees and stands under severe drought：a review of ecophysiological responses，adaptation processes and long-term consequences［J］.Annals of Forest Science，2006，63（6）：625-644.
5	FICHOT R， LAURANS F， MONCLUS R，et al.Xylem anatomy correlates with gas exchange，water-use efficiency and growth performance under contrasting water regimes：evidence from Populus deltoides × Populus nigra hybrids［J］.Tree Physiology，2009，29（12）：1537-1549.
6	LIMOUSIN J M， LONGEPIERRE D，HUC R，et al.Change in hydraulic traits of mediterranean Quercus ilex subjected to long-term throughfall exclusion［J］.Tree Physiology，2010，30（8）：1026-1036.
7	VENTURAS M D， SPERRY J S， HACKE U G.Plant xylem hydraulics：What we understand，current research，and future challenges［J］.Journal of Integrative Plant Biology，2017，59（6）：356-389.
8	TONG S F， WANG Y B， CHEN N N，et al.Ptonf-yc9-srmt-ptord26 module regulates the high saline tolerance of a triploid poplar［J］.Genome Biology，2022，23（1）：148.
9	LI S， LIN Y C J， WANG P Y. et al.The AREB1 transcription factor influences histone acetylation to regulate drought responses and tolerance in Populus trichocarpa ［J］.The Plant Cell，2019，31（3）：663-686.
10	杜兆伟，郑唐春，李爽，等.小黑杨快繁与再生体系的优化［J］.植物研究，2015，35（6）：904-907.
	DU Z W， ZHENG T C， LI S，et al.Rapid propagation and regeneration system of Populus simonii×Populus nigra ［J］.Bulletin of Botanical Research，2015，35（6）：904-907.
11	李晶，王福森，李树森，等.几个杨树新品系抗寒性测定试验［J］.防护林科技，2012（1）：59-61.
	LI J， WANG F S， LI S S，et al.Cold-resistance determination for several new strains of poplar［J］.Protection Forest Science and Technology，2012（1）：59-61.
12	何旭，高源，张群野，等.白城小黑杨遗传转化体系建立及其应用［J］.植物研究，2023，43（5）：667-678.
	HE X， GAO Y， ZHANG Q Y，et al.Establishment and application of genetic transformation system for Populus simonii×P.nigra ‘Baicheng’［J］.Bulletin of Botanical Research，2023，43（5）：667-678.
13	ILYAS M， NISAR M， KHAN N，et al.Drought tolerance strategies in plants：a mechanistic approach［J］.Journal of Plant Growth Regulation，2020，40（3）：926-944.
14	PASCUAL M B， DE LA TORRE F， CAÑAS R A，et al.NAC transcription factors in woody plants［J］.Progress in Botany，2018，80：195-222.
15	HAN K J， ZHAO Y， SUN Y H，et al.NACs，generalist in plant life［J］.Plant Biotechnology Journal，2023，21（12）：2433-2457.
16	NURUZZAMAN M， MANIMEKALAI R， SHARONI A M，et al.Genome-wide analysis of NAC transcription factor family in rice［J］.Gene，2010，465（1/2）：30-44.
17	JEONG J S， KIM Y S， BAEK K H，et al.Root-specific expression of OsNAC10 improves drought tolerance and grain yield in rice under field drought conditions［J］.Plant Physiology，2010，153（1）：185-197.
18	CHEN D D， CHAI S C， MCINTYRE C L，et al.Overexpression of a predominantly root-expressed NAC transcription factor in wheat roots enhances root length，biomass and drought tolerance［J］.Plant Cell Reports，2018，37（2）：225-237.
19	YANG C F， HUANG Y Z， LV W H，et al. GmNAC8 acts as a positive regulator in soybean drought stress［J］.Plant Science，2020，293：110442.
20	WANG H L， YANG Q， TAN S Y，et al.Regulation of cytokinin biosynthesis using PtRD26_pro-IPT module improves drought tolerance through PtARR10-PtYUC4/5-mediated reactive oxygen species removal in Populus ［J］.Journal of Integrative Plant Biology，2022，64（3）：771-786.
21	CHEN Z H， PENG Z X， LIU S Q，et al.Overexpression of PeNAC122 gene promotes wood formation and tolerance to osmotic stress in poplars［J］.Physiologia Plantarum，2022，174（4）：e13751.
22	李开隆，杨传平，刘桂丰.黑龙江省杨树遗传育种研究进展［J］.东北林业大学学报，2003，31（4）：45-48.
	LI K L， YANG C P， LIU G F.Research Progresses of genetics and breeding of Populus in Heilongjiang province［J］.Journal of Northeast Forestry University，2003，31（4）：45-48.
23	吴丽丽，高福玲，王雷，等.杨树幼茎特异表达基因及PsnLAC基因的克隆［J］.东北林业大学学报，2011，39（4）：5-7.
	WU L L， GAO F L， WANG L，et al.Specific gene expression in stem of Populus simonii×P.nigra and cloning of PsnLAC gene［J］.Journal of Northeast Forestry University，2011，39（4）：5-7.
24	王遂，刘梦然，黄海娇，等.转TaLEA基因小黑杨抗寒株系的筛选［J］.东北林业大学学报，2011，39（9）：5-7.
	WANG S， LIU M R， HUANG H J，et al.Selection of cold resistant strains from TaLEA gene transferred Populus simonii×P.nigra ［J］.Journal of Northeast Forestry University，2011，39（9）：5-7.
25	WANG S J， FAN Y， DU S H. et al.PtaERF194 inhibits plant growth and enhances drought tolerance in poplar［J］.Tree Physiology，2022，42（8）：1678-1692.
26	RODRIGUEZ-ZACCARO F D， GROOVER A.Wood and water：How trees modify wood development to cope with drought［J］.Plants，People，Planet，2019，1（4）：346-355.
27	ECKERT C， SHARMIN S， KOGEL A，et al.What makes the wood？Exploring the molecular mechanisms of xylem acclimation in hardwoods to an ever-changing environment［J］.Forests，2019，10（4）：358.
28	BARIGAH T S， CHARRIER O， DOURIS M，et al.Water stress-induced xylem hydraulic failure is a causal factor of tree mortality in beech and poplar［J］.Annals of Botany，2013，112（7）：1431-1437.
29	LAUDER J D.From the cell to the stand：trait-based approaches to understanding forest response to climate cha-nge［M］.Merced：University of California，2020：101-113.
30	BANG S W， LEE D K， JUNG H，et al.Overexpression of OsTF1L，a rice HD-Zip transcription factor，promotes lignin biosynthesis and stomatal closure that improves drought tolerance［J］.Plant Biotechnology Journal，2019，17（1）：118-131.
31	MÉNARD D， BLASCHEK L， KRIECHBAUM K，et al.Plant biomechanics and resilience to environmental changes are controlled by specific lignin chemistries in each vascular cell type and morphotype［J］.The Plant Cell，2022，34（12）：4877-4896.
32	SAINI R， KAUR A， SAINI J K，et al.Trends in lignin biotransformations for bio-based products and energy applications［J］.BioEnergy Research，2023，16（1）：88-104.
33	ZAHRA N， HAFEEZ M B， KAUSAR A，et al.Plant photosynthetic responses under drought stress：effects and management［J］.Journal of Agronomy and Crop Science，2023，209（5）：651-672.

引物名称 Primer name	引物序列（5′→3′） Sequences（5′→3′）	用途 Application
PsnNAC007-F	CACCATGAAAGGAAATAGATCAGCAG	PsnNAC007基因克隆 PsnNAC007 gene cloning
PsnNAC007-R	GCACACATCAAGAACCCTATT	PsnNAC007基因克隆 PsnNAC007 gene cloning
PsnNAC007-XbaⅠ-F	CTAGTCTAGAATGAAAGGAAATAGATCAGCAG	pBI121-35S-PsnNAC007载体构建 pBI121-35S-PsnNAC007 vector construction
PsnNAC007-SacⅠ-R	CTAGGAGCTCGCACACATCAAGAACCCTATT
35S-F	CCCACTATCCTTCGCAAGACC	转基因鉴定 Transgenic identification
PsnNAC007-middle-R	CGACCAATGGGTTTATCTGCTC	转基因鉴定 Transgenic identification
RT-PsnActin-F	GGATATTCAGCCCCTTGTCTG-	相对表达量检测引物 Relative expression level detection primers
RT-PsnActin-R	TCGTCACCAACATAAGCATCC
RT-PsnNAC007-F	ATGAAAGGAAATAGATCAGCAGAT
RT-PsnNAC007-R	ATTGGCCTCCACATTTCTTAAGC

参数 Parameter	野生型 Wild-type/%	PsnNAC007过表达植株 OE-PsnNAC007/%
不可溶性木质素 Acid-insoluble lignin	20.32±0.23	22.43±0.28**
可溶性木质素 Acid-soluble lignin	2.96±0.07	2.64±0.03**
总木质素Total lignin	23.29±0.29	25.07±0.30**
葡萄糖Glucose	53.37±0.17	53.63±0.19
木糖Xylose	11.39±0.26	11.59±0.40
半乳糖Galactose	1.34±0.05	1.28±0.03
阿拉伯糖Arabinose	2.30±0.13	2.29±0.05
总碳水化合物 Total carbohydrate	68.41±0.38	68.79±0.48

[1]	Dongyue WANG, Ruyue WANG, Maotong SUN, Cuishuang LIU, Jihong LI. Establishment of Genetic Transformation System for ‘Populus leucopyramidalis 1’ and Transformation of Insect Resistance Gene [J]. Bulletin of Botanical Research, 2024, 44(3): 361-369.
[2]	Xu HE, Yuan GAO, Qunye ZHANG, Chenguang ZHOU, Wei LI, Shuang LI. Establishment and Application of Genetic Transformation System for Populus simonii×P. nigra ‘Baicheng’ [J]. Bulletin of Botanical Research, 2023, 43(5): 667-678.
[3]	Yu SUN, Yiteng ZHANG, Huihui CHENG. The Function of Salt and Alkaline Tolerance of WRKY42 Gene in Amorpha fruticosa [J]. Bulletin of Botanical Research, 2023, 43(4): 612-621.
[4]	Zhanmin ZHENG, Yubing SHANG, Guangbo ZHOU, Di XIAO, Yi LIU, Xiangling YOU. Genetic Transformation and Function Analysis of PsnHB13 and PsnHB15 of Populus simonii × Populus nigra [J]. Bulletin of Botanical Research, 2023, 43(3): 340-350.
[5]	Shixian LIAO, Yuting WANG, Liben DONG, Yongmei GU, Fenglin JIA, Tingbo JIANG, Boru ZHOU. Function Analysis of the Transcription Factor PsnbZIP1 of Populus simonii×P. nigra in Response to Salt Stress [J]. Bulletin of Botanical Research, 2023, 43(2): 288-299.
[6]	Senyao LIU, Fenglin JIA, Qing GUO, Gaofeng FAN, Boru ZHOU, Tingbo JIANG. Response Analysis of Transcription Factor PsnbHLH162 Gene in Populus simonii × P. nigra under Salt Stress and Low Temperature Stress [J]. Bulletin of Botanical Research, 2023, 43(2): 300-310.
[7]	Jinxia DU, Tingting SHEN, Haoran WANG, Yiping LIN, Huiyu LI, Lianfei ZHANG. Construction of Suppression Expression Vector and Genetic Transformation of BpSPL9 gene from Betula platyphylla [J]. Bulletin of Botanical Research, 2023, 43(1): 30-35.
[8]	Ting QIAN, Fan ZHAO, Yujie ZHANG, Xueli LI, Kun SUN, Hui ZHANG. Study on Adaptive Differentiation of Transcription Factor bHLH94 Gene to Altitude in Hippophae neurocarpa and H.tibetana [J]. Bulletin of Botanical Research, 2022, 42(6): 976-985.
[9]	Denggao LI, Rui LIN, Qinghui MU, Na ZHOU, Yanru ZHANG, Wei BAI. Cloning and Functional Analysis of StNPR4 gene in Solanum tuberosum [J]. Bulletin of Botanical Research, 2022, 42(5): 821-829.
[10]	Sisi CHEN, Muhong XIE, Maokai CUI, Wenkai LI, Zhenggang XU, Caixia JIA, Guiyan YANG. Identification of Broussonetia papyrifera Transcription Factor BpbZIP1 and Analysis of Its Response to Cadmium Stress [J]. Bulletin of Botanical Research, 2022, 42(3): 394-402.
[11]	Bin WEI, Yi LI, Shiping SU. The Effect of Exogenous Proline on the Stomata of Nitraria tangutorum Leaves under Natural Drought [J]. Bulletin of Botanical Research, 2022, 42(3): 492-501.
[12]	Qi Li, Siyu Yan, Su Chen. Genetic Transformation of BpERF98 Gene and Abiotic Stress Response of Transgenic Plant in Betula platyphylla [J]. Bulletin of Botanical Research, 2022, 42(1): 93-103.
[13]	Lian-Bin HAN, Qing GUO, Kai ZHAO, Ting-Bo JIANG, Bo-Ru ZHOU, Li LI. Cloning and Expression Analysis of HD-Zip Transcription Factor PsnHB63 in Populus simonii × P. nigra [J]. Bulletin of Botanical Research, 2021, 41(6): 1006-1014.
[14]	Feng HE, Hong-Yan DU, Pan-Feng LIU, Lu WANG, Jun QING, Lan-Ying DU. Effects of Drought Stress on Leaf Structure of Eucommia ulmoides [J]. Bulletin of Botanical Research, 2021, 41(6): 947-956.
[15]	Shuang-Hui TIAN, He CHENG, Yang ZHANG, Cong LIU, De-An XIA, Zhi-Gang WEI. Genome-wide Identification and Expressional Analysis of Carotenoid Cleavage Dioxygenases(CCD) Gene Family in Populus trichocarpa under Drought and Salt Stress [J]. Bulletin of Botanical Research, 2021, 41(6): 993-1005.