Growth-promoting Mechanism of Trichoderma harzianum T9131 on Astragalus membranaceus Seedlings

doi:10.13560/j.cnki.biotech.bull.1985.2025-0377

Abstract

Abstract:

Objective To explore the regulatory mechanism of Trichoderma harzianum T9131 on the growth promotion of Astragalus membranaceus seedlings and provide a theoretical basis for T9131 to increase the yield of A. membranaceus. Method The A. membranaceus seedlings grown for 30 d were treated by root irrigation with T. harzianum T9131 spores suspension. Samples were taken from the roots of the A. membranaceus seedlings at 0, 24 h, 48 h and 45 d after root irrigation, and the plant height, root length and fresh weight of the A. membranaceus seedlings at 45 d were determined. Transcriptome libraries were constructed at three time points of 0, 24 and 48 h by RNA-Seq. Differentially expressed genes (DEGs) in the three comparison groups (T_24 h vs T_0 h, T_48 h vs T_0 h, and T_48 h vs T_24 h) were obtained by using DESeq2 software. Venn analysis was conducted on the three groups of DEGs to clarify that the three groups had a total of DEGs. KEGG enrichment analysis was performed on the common DEGs to screen the growth-promoting related genes, and the expression of growth-promoting-related genes in A. membranaceus seedlings in the early stage after 45 d of induction with T9131 and induction with exogenous hormone ABA was analyzed by RT-qPCR. The contents of 15 hormones in the root samples at 0 h, 48 h and 45 d were determined by the LC-MS/MS method. Result T9131 could significantly promote the plant height and fresh weight of A. membranaceus seedlings.Transcriptome analysis of A. membranaceus seedlings induced by T9131 indicated that there were 149 differentially expressed genes in the three comparison groups. KEGG enrichment pathway analysis showed that the pathways related to growth promotion mainly included terpenoid backbone biosynthesis, carotenoid biosynthesis, diterpenoid biosynthesis, cyanoamino acid metabolism, zeatin biosynthesis, and plant hormone signal transduction. The gene types enriched in the pathway include HMGR, CYP707A2, GA2ox2, E3.2.1.21, UGT73C6, PYL4, GH3.1, and JAZ, totaling 14 genes. Except for the gene UGT73C6 (DN24891_c1_g1) down-regulated by T9131 induction, the remaining genes were all up-regulated in expression. Fourteen genes induced A. membranaceus seedlings in T9131 at 45 d, seven genes including HMGR (DN10463_c0_g2, DN989_c0_g1, DN113450_c0_g1, DN989_c2_g1), CYP707A2 and JAZ were significantly up-regulated, while the remaining genes were down-regulated. Analysis of the expressions of the 14 genes induced by ABA showed that the expressions of 6 genes, including HMGR (DN10463_c0_g2, DN989_c0_g1, DN113450_c0_g1, DN989_c2_g1), CYP707A2 (DN397_c0_g2), and E3.2.1.21 (DN2073_c0_g1), were significantly down-regulated by ABA induction, while the expressions of the remaining genes were up-regulated. Hormone determination indicated that T9131 significantly reduced the content of ABA. Conclusion T. harzianum T9131 may promote the growth of A. membranaceus by regulating the up-regulated expressions of genes related to the growth promotion of A. membranaceus, such as HMGR, CYP707A2 and JAZ, and by reducing the content of ABA. Exogenous ABA can reduce the expressions of HMGR and CYP707A2.

Key words: Trichoderma harzianum, Astragalus membranaceus, growth index, growth-promoting regulation, transcriptome, LC-MS/MS, hormone content, expression analysis

BAI Yu-guo, LI Wan-di, LIANG Jian-ping, SHI Zhi-yong, LU Geng-long, LIU Hong-jun, NIU Jing-ping. Growth-promoting Mechanism of Trichoderma harzianum T9131 on Astragalus membranaceus Seedlings[J]. Biotechnology Bulletin, 2025, 41(8): 175-185.

Figures/Tables 12

References 41

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基因名称 Gene name	基因序号 Gene_ID	引物序列 Sequence （5′‒3′）
HMGR	DN10463_c0_g2	F： CACTGCACGGACAACTTCC	R： ATTGCCGACCAGAAACCC
	DN989_c0_g1	F： GGATTCCCAATCAGTGCC	R： ATTCGTCTTCGTCGTAGTAGGT
	DN113450_c0_g1	F： AGTGGCTGGTTCTGTTCTCG	R： TGTGGCGTTATCGTAGGGA
	DN52213_c0_g1	F： AAGGATTCCCAATCAGTGCC	R： CGAAGAAGATGACGAGGTAGATG
	DN989_c2_g1	F： TTTCCCTTCTCGGATTTCA	R： AAGACCGCGTTGGTCACGT
CYP707A2	DN397_c0_g1	F： CTCATAGCCTTGTGGAATAGTGT	R： AGAGCGTTTGAAGCAGTGTTA
CYP707A2	DN397_c0_g2	F： ACTGGCAGTAGTGTCACGAGC	R： CAAGGAGGTGTATGAAGCAAAA
GA2ox2	DN7261_c0_g1	F： TGAACCACCGTGTACCAAAG	R： ATTCCAGGTCACCCATATCAC
E3.2.1.21	DN19827_c0_g1	F： AGCAAGAAGTTGGTAGTGGGTA	R： GATTAGAAGATGAGTATGGTGGC
E3.2.1.21	DN2073_c0_g1	F： AAGAACATCGACCTGGTGC	R： ACTTTAGGGATTATGCGGAAC
UGT73C6	DN24891_c1_g1	F： CCAGGGTAGTGGGTAGGAGT	R： GAAGACGTACAAGCGGGAG
PYL4	DN9921_c0_g2	F： TAACTAACGCCATAGCAAACG	R： ACTTCACGAAGCGATCCAA
GH3.1	DN11968_c0_g1	F： TCCATTAGGTCCACCAGCAT	R： TGTACGGCATCGGCATTT
JAZ	DN64262_c0_g1	F： CCAGGAACCGATGAAGTGA	R： ACCCTACAAAGGCAGCAGA
Am18S		F： TGCAGAATCCCGTGAACCATC	R： AGGCATCGGGCAACGATATG

基因名称 Gene name	基因序号 Gene_ID	引物序列 Sequence （5′‒3′）
HMGR	DN10463_c0_g2	F： CACTGCACGGACAACTTCC	R： ATTGCCGACCAGAAACCC
	DN989_c0_g1	F： GGATTCCCAATCAGTGCC	R： ATTCGTCTTCGTCGTAGTAGGT
	DN113450_c0_g1	F： AGTGGCTGGTTCTGTTCTCG	R： TGTGGCGTTATCGTAGGGA
	DN52213_c0_g1	F： AAGGATTCCCAATCAGTGCC	R： CGAAGAAGATGACGAGGTAGATG
	DN989_c2_g1	F： TTTCCCTTCTCGGATTTCA	R： AAGACCGCGTTGGTCACGT
CYP707A2	DN397_c0_g1	F： CTCATAGCCTTGTGGAATAGTGT	R： AGAGCGTTTGAAGCAGTGTTA
CYP707A2	DN397_c0_g2	F： ACTGGCAGTAGTGTCACGAGC	R： CAAGGAGGTGTATGAAGCAAAA
GA2ox2	DN7261_c0_g1	F： TGAACCACCGTGTACCAAAG	R： ATTCCAGGTCACCCATATCAC
E3.2.1.21	DN19827_c0_g1	F： AGCAAGAAGTTGGTAGTGGGTA	R： GATTAGAAGATGAGTATGGTGGC
E3.2.1.21	DN2073_c0_g1	F： AAGAACATCGACCTGGTGC	R： ACTTTAGGGATTATGCGGAAC
UGT73C6	DN24891_c1_g1	F： CCAGGGTAGTGGGTAGGAGT	R： GAAGACGTACAAGCGGGAG
PYL4	DN9921_c0_g2	F： TAACTAACGCCATAGCAAACG	R： ACTTCACGAAGCGATCCAA
GH3.1	DN11968_c0_g1	F： TCCATTAGGTCCACCAGCAT	R： TGTACGGCATCGGCATTT
JAZ	DN64262_c0_g1	F： CCAGGAACCGATGAAGTGA	R： ACCCTACAAAGGCAGCAGA
Am18S		F： TGCAGAATCCCGTGAACCATC	R： AGGCATCGGGCAACGATATG

样品名称 Sample	过滤后的reads数 Clean reads	过滤后的总数据量 Clean bases	Q20 （%）	Q30 （%）	GC含量比例 GC content （%）
T_0 h_1	49 844 534	7 453 178 401	98.27	94.75	42.15
T_0 h_2	41 238 544	6 167 259 149	98.19	94.54	42.31
T_0 h_3	53 377 736	7 964 860 750	98.31	94.88	42.28
T_24 h_1	42 676 268	6 312 001 975	98.27	94.77	42.34
T_24 h_2	47 351 528	7 061 996 999	98.19	94.55	42.45
T_24 h_3	51 754 072	7 733 095 911	98.26	94.75	42.63
T_48 h_1	43 661 406	6 521 807 743	98.02	94.06	42.41
T_48 h_2	50 643 772	7 548 606 160	98.30	94.86	42.29
T_48 h_3	47 716 402	7 100 483 345	98.29	94.82	42.56

样品名称 Sample	过滤后的reads数 Clean reads	过滤后的总数据量 Clean bases	Q20 （%）	Q30 （%）	GC含量比例 GC content （%）
T_0 h_1	49 844 534	7 453 178 401	98.27	94.75	42.15
T_0 h_2	41 238 544	6 167 259 149	98.19	94.54	42.31
T_0 h_3	53 377 736	7 964 860 750	98.31	94.88	42.28
T_24 h_1	42 676 268	6 312 001 975	98.27	94.77	42.34
T_24 h_2	47 351 528	7 061 996 999	98.19	94.55	42.45
T_24 h_3	51 754 072	7 733 095 911	98.26	94.75	42.63
T_48 h_1	43 661 406	6 521 807 743	98.02	94.06	42.41
T_48 h_2	50 643 772	7 548 606 160	98.30	94.86	42.29
T_48 h_3	47 716 402	7 100 483 345	98.29	94.82	42.56

KEGG通路 KEGG pathway	基因名称 Gene name	基因功能描述 Gene function description	基因ID Gene_ID	T_24 h vs T_0 h （log₂FC）	T_48 h vs T_0 h （log₂FC）	T_48 h vs T_24 h （log₂FC）
萜类骨架生物合成 Terpenoid backbone biosynthesis （map00900）	HMGR	3-羟基-3-甲基戊二酰辅酶A还原酶	DN10463_c0_g2	2.129 9	3.611 3	1.457 1
			DN989_c0_g1	1.904 1	3.338 9	1.409 5
			DN113450_c0_g1	1.599 3	3.385 2	1.771 3
			DN52213_c0_g1	2.369 7	3.898 2	1.497 5
			DN989_c2_g1	2.061 1	3.914 2	1.825 3
类胡萝卜素生物合成 Carotenoid biosynthesis （map00906）	CYP707A2	ABA 8'-羟化酶编码基因	DN397_c0_g2	1.825 3	4.789 2	1.597 6
类胡萝卜素生物合成 Carotenoid biosynthesis （map00906）	CYP707A2	ABA 8'-羟化酶编码基因	DN397_c0_g1	2.256 5	3.766 6	1.482 3
二萜生物合成 Diterpenoid biosynthesis （map00904）	GA2ox2	赤霉素2-氧化酶	DN7261_c0_g1	1.871 7	4.344 5	2.441 8
氰基氨基酸代谢 Cyanoamino acid metabolism （map00460）	E3.2.1.21	β-葡萄糖苷酶	DN19827_c0_g1	2.245 8	3.315 2	1.043 4
氰基氨基酸代谢 Cyanoamino acid metabolism （map00460）	E3.2.1.21	β-葡萄糖苷酶	DN2073_c0_g1	2.346 5	3.672 8	1.298 6
玉米素生物合成 Zeatin biosynthesis （map00908）	UGT73C6	UDP-糖基转移酶	DN24891_c1_g1	-1.356 9	-2.907 5	-1.565 5
植物激素信号转导 Plant hormone signal transduction （map04075）	PYL4	ABA受体	DN9921_c0_g2	2.107 7	3.822 5	1.698 2
	GH3.1	吲哚-3-乙酸酰胺合成酶	DN11968_c0_g1	1.889 8	3.488 6	1.571 7
	JAZ	TIFY10A蛋白	DN64262_c0_g1	2.062 9	3.588 1	1.499 8