Characterization of IpSAD gene family in Idesia polycarpa and functional identification

doi:10.12302/j.issn.1000-2006.202207034

JOURNAL OF NANJING FORESTRY UNIVERSITY ›› 2022, Vol. 46 ›› Issue (6): 261-270.doi: 10.12302/j.issn.1000-2006.202207034

Special Issue: 南京林业大学120周年校庆特刊

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Characterization of IpSAD gene family in Idesia polycarpa and functional identification

GONG Jue¹(), BI Hao¹, YANG Wenlu¹, WANG Dongming², WANG Lehui³, JIANG Yuanzhong¹, MA Tao¹^,^*()

1. School of Life Sciences, Sichuan University, Chengdu 610064, China
2. Ziyang Yanjiang District Agricultural and Rural Bureau of Sichuan Province, Ziyang 641300, China
3. Sichuan Dantong Technology Co., Ltd., Chengdu 610014, China

Received:2022-07-21 Revised:2022-09-16 Online:2022-11-30 Published:2022-11-24
Contact: MA Tao E-mail:455948475@qq.com;13519669951@139.com

Abstract

Abstract:

【Objective】Stearoyl-ACP Δ⁹ desaturase (SAD) is a key enzyme that determines the composition of fatty acids. The whole-genome identification of Idesia polycarpa SAD genes can provide targets to improve fatty acid compositions by genetic methods.【Method】HMM and Blastp were used to identify IpSAD genes in the I. polycarpa genome. The gene structures, phylogenetic relationship, conserved motifs, physicochemical characters of IpSADs, and cis-regulatory element in these gene promoters were analyzed and predicted by MEGA X, MEME and PlantCare online software, respectively. The MCSCANX software was used to reveal the common linear relationship of SAD genes in I. polycarpa and P. trichocarpa. The tissue expression pattern of IpSAD genes was constructed according to RNA-seq, and the function of these genes was identified by a virus-induced gene silencing (VIGS) system.【Result】(1)A total of seven IpSAD genes (IpSAD1-7) were identified in the I. polycarpa genome, and these family members shared high sequence similarity and conserved domains. (2)Aphylogenetic tree indicated that IpSAD members were divided into three clades, which were consistent with the classification of the homologs in Arabidopsis thaliana. In particular,IpSAD2, IpSAD3 and IpSAD4 were closer to AtSSI2, which is the key enzyme of oleic acid biosynthesis. (3)The cis-regulatory elements in the promoters of IpSAD genes included elements of light and abscisic acid responses. (4)These IpSAD genes were divided into three categories according to their tissue expression patterns. The IpSAD genes in Classes Ⅰ and Ⅱ were expressed with a low level in most tissues, whereas Class Ⅲ genes showed a high level in all tested tissues. (5) Silencing IpSAD genes using VIGS significantly reduced the oleic acid content of leaves.Remarkably, the oleic acid content in IpSAD3 silenced leaves was decreased by up to 76%.【Conclusion】The characteristics of each IpSAD gene was clarified, and their functions in oleic acid synthesis were determined. Our findings can aid in the investigation of the biosynthesis mechanisms of oleic acid and total oil in I. polycarpa.

Key words: Idesia polycarpa, stearoyl-acyl carrier protein Δ⁹ desaturase (SAD), oleic acid, virus induced gene silencing(VIGS)

CLC Number:

S722.37
Q78

GONG Jue, BI Hao, YANG Wenlu, WANG Dongming, WANG Lehui, JIANG Yuanzhong, MA Tao. Characterization of IpSAD gene family in Idesia polycarpa and functional identification[J]. JOURNAL OF NANJING FORESTRY UNIVERSITY, 2022, 46(6): 261-270.

Figures/Tables 9

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引物名称 primer name	序列 sequence	目的 purpose
SAD2-F	ATTCTCTAGAAGGCCTCCATGG GAAGCCAAATTACAGTGAGG
SAD2-R	CGGGACATGCCCGGGCCTCGAG GCTCTTCCACCAATGGCTAG
SAD3-F	ATTCTCTAGAAGGCCTCCATGGCGACTTGTTCATGCACCTCC
SAD3-R	CGGGACATGCCCGGGCCTCGAG ATGGCTCTCAAGCTCAATCC
SAD4-F	ATTCTCTAGAAGGCCTCCATGGATGAAACATGCAAAGATCCA
SAD4-R	CGGGACATGCCCGGGCCTCGAG AGCTCAATCCTTTTGTCTCT	VIGS
SAD5-F	CATG CCATGG TGGTGGAAATGGAATTCAAA
SAD5-R	CC CTCGAG CCTTCCCATCAATTCACAAC
SAD6-F	CATG CCATGG AGTGATGCACCAGTCTCGTC
SAD6-R	CC CTCGAG CCATGGAGAATTGGGCTAAG
SAD7-F	ATTCTCTAGAAGGCCTCCATGGCAGGGCTTCTTCAGTAATCA
SAD7-R	CGGGACATGCCCGGGCCTCGAG GATAACCTCTTGACGCTCCT
Q-IpSAD2-F	ATTTGGACAAGGGCATGGAC
Q-IpSAD2-R	ATGAACCCCAGGTAGGGACT
Q-IpSAD3-F	TCACATCATGGGCAATCTGG
Q-IpSAD3-R	TGGACTGTTCTCTGTTCCCG
Q-IpSAD4-F	AGCGTCTTGGAGTCTACACA	qRT-PCR
Q-IpSAD4-R	GAGGTAACCCGCAGACATAA
Q-IpSAD5-F	ACCTTCCTTCTCAATCTCGC
Q-IpSAD5-R	TTGTGGAGGCATAGAATGGG
Q-IpSAD6-F	ATGGCTCTCAAGCTGAACTC
Q-IpSAD6-R	GTGCCATGGAATGTGTCACT
Q-IpSAD7-F	TCTTATCGGGTCAGGAATGG
Q-IpSAD7-R	TCCACAATCTTGGTGTAGGC
UBQ-F	TTGACGGGAAAGACCATAAC
UBQ-R	CTTTCTCTTCTTGGCTCCAC

基因名 gene name	基因编号 gene ID	蛋白质信息 calculated protein information			理论等电点 theoretical pI
基因名 gene name	基因编号 gene ID	长度/aa length	分子质量/ku MW	亚细胞定位 subcellular localization	理论等电点 theoretical pI
IpSAD1	Idesia.03G00684	399	45.29	叶绿体	6.47
IpSAD2	Idesia.05G01258	396	45.29	叶绿体	5.99
IpSAD3	Idesia.07G00300	396	45.16	叶绿体	5.99
IpSAD4	Idesia.07G00301	398	45.23	叶绿体	6.11
IpSAD5	Idesia.18G01639	397	45.26	叶绿体	6.11
IpSAD6	Idesia.18G01640	386	44.47	叶绿体	6.64
IpSAD7	Idesia.20G00641	333	38.28	叶绿体	5.15

物种 species	来源 accession
拟南芥 Arabidopsis thaliana	At1g43800、At2g43710、At3g02610、At3g02620、At3g02630、At5g16230、At5g16240
毛果杨 Populus trichocarpa	Potri.010G179400、Potri.010G179500、Potri.013G127300、Potri.T034100、Potri.005G187600、Potri.008G077900
可可 Theobroma cacao	XP_007032081、XP_017974119、XP_017974120、XP_017974175、XP_017975512、XP_017977433、XP_017980831
油菜 Brassica nupus	BnaA01T0414100ZS、BnaA03T0212900ZS、BnaA03T0288900ZS,BnaA05T0039900ZS、BnaA10T0202500ZS、BnaA10T0202600ZS、BnaC01T0501000ZS、BnaC03T0251400ZS、BnaC03T0347000ZS,BnaC04T0043700ZS、BnaC09T0500900ZS、BnaC09T0501100ZS

Characterization of IpSAD gene family in Idesia polycarpa and functional identification

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