Study on the Regulatory Mechanism of SlPL Gene Affecting Tomato Fruit Cracking

doi:10.16420/j.issn.0513-353x.2023-0816

Acta Horticulturae Sinica ›› 2024, Vol. 51 ›› Issue (2): 295-308.doi: 10.16420/j.issn.0513-353x.2023-0816

• Genetic&Breeding?Germplasm Resources?Molecular Biology • Previous Articles Next Articles

Study on the Regulatory Mechanism of SlPL Gene Affecting Tomato Fruit Cracking

ZHONG Zhaojiang¹, WU Zhen¹, ZHOU Rong¹, ZHU Weimin², YANG Xuedong², YU Xiaowei¹, XU Yan¹, GAO Yangyang¹, JIANG Fangling¹^,^*()

¹ Department of Horticulture，Nanjing Agricultural University，Nanjing 210095，China
² Institute of Horticulture，Shanghai Academy of Agricultural Sciences，Shanghai 201403，China

Received:2023-10-18 Revised:2023-12-10 Online:2024-02-25 Published:2024-02-26
Contact: JIANG Fangling

Abstract

Abstract:

Tomato with highly significant differences in fruit cracking rate were used as the materials to analyze the expression differences of the SlPL（Solyc03g111690）gene，which was further functionally validated using genetic transformation. The results showed that the expression of SlPL was significantly higher in crack-susceptible tomato‘NT189’than in crack-resistant tomato‘NT91’；The expression of SlPL in tomato fruit was significantly higher than that in other organs such as roots，stems，leaves and flowers，and the expression was higher in the break ripen and red ripen stages of the fruit. Fruit cracking was further induced by irrigation and ABA treatments，and it was found that the expression of SlPL was overall significantly higher in the fruit of crack-susceptible materials than that of crack-resistant ones in the same treatment period. SlPL overexpression（OEPL）and knockout（pl）lines were then obtained by genetic transformation. OEPL was more susceptible to fruit cracking and had significantly lower fruit firmness and pl had higher fruit firmness than the wild type. OEPL fruit had significantly lower pro-pectin content than the wild type，and significantly higher water-soluble pectin content than the wild type，while pl fruit had significantly higher pro-pectin and total pectin content than the wild type. Pectin cleavage enzyme activity was significantly higher in OEPL fruits than in wild type，and pectin cleavage enzyme activity was significantly lower in pl fruits than in wild type. Gene expression analysis revealed that the relative expression of cell wall metabolism-related genes SlPG2，SlPME2.1，SlCel2，SlGH9C5，and ethylene synthesis pathway-related genes SlACS4 and SlACO1 was significantly higher in OEPL fruits than in wild type，and the opposite was true for pl fruits. pl fruits showed a significantly higher relative expression of the ethylene-responsive factor SlERF2 than in wild type. Observations of the pericarp microstructure revealed that the cells of the epidermal layer and thin-walled cells were more sparsely arranged in OEPL compared with wild type，and those cells were tightly arranged in pl.

Key words: tomato, fruit cracking, SlPL, functional identification

ZHONG Zhaojiang, WU Zhen, ZHOU Rong, ZHU Weimin, YANG Xuedong, YU Xiaowei, XU Yan, GAO Yangyang, JIANG Fangling. Study on the Regulatory Mechanism of SlPL Gene Affecting Tomato Fruit Cracking[J]. Acta Horticulturae Sinica, 2024, 51(2): 295-308.

Figures/Tables 10

References 40

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用途 Purpose	名称 Name	序列（5′-3′） Sequence
基因克隆引物 Primer for genes clone	pbi121-SlPL-F	gagaacacgggggactctagaATGGGCACTTCCTCTGTTTTTC
基因克隆引物 Primer for genes clone	pbi121-SlPL-R	ataagggactgaccacccgggGCAACGAGAACCCTTTTTACAGTT
扩增引物 Amplification primers	CRISPR-SlPL-F	ACGAAGGAACAAGGGACCCA
扩增引物 Amplification primers	CRISPR-SlPL-R	CGGGTGGAGAACTGTTTCGG
测序引物 Sequencing primers	CRISPR-SlPL-F	CAGATAGGAACAAGGAGT
测序引物 Sequencing primers	CRISPR-SlPL-R	GGTGGAAATGCTATGGTG
	35S-F	TCCCACTGAATCAAAGGC
qPCR引物 Primer for qPCR	Actin-F	CTCTACATACTTGAGAGGTGCC
qPCR引物 Primer for qPCR	Actin-R	AGACGAGGAGAAAACATCACAA
	SlPL-F	GGAAATCCAATCGACGATTGTT
	SlPL-R	CGTTTTTCCCAAATCCAATTGC
	SlCel2-F	TATCAAATGGCGTAGAGACTCC
	SlCel2-R	ACATTATCTCCGGCATCGTAAT
	SlPG2-F	CCAAAGGAATAGTATTCTCCTTCTC
	SlPG2-R	GTTTTTCCATCACCCTTAGCTC
	SlPME2.1-F	ATGCTACCATCATTACTGGGAG
	SlPME2.1-R	GATAAGCATCGATACGACAACG
	SlGH9C2-F	CCAGAAGTACTACCGATCTGTC
	SlGH9C2-R	TCAGTCATAGACCAACCAGTTC
	SlACO1-F	CATACAGACGCAGGAGGCATCA
	SlACO1-R	TAGAGTGGCGCATGGGAGGAA
	SlACS4-F	CTCCTCAAATGGGGAGTACG
	SlACS4-R	TTTTGTTTGCTCGCACTACG
	SlERF2-F	CGACCTATGGCCGACTGATT
	SlERF2-R	GGTCACGAATTTCAGCAGCC

用途 Purpose	名称 Name	序列（5′-3′） Sequence
基因克隆引物 Primer for genes clone	pbi121-SlPL-F	gagaacacgggggactctagaATGGGCACTTCCTCTGTTTTTC
基因克隆引物 Primer for genes clone	pbi121-SlPL-R	ataagggactgaccacccgggGCAACGAGAACCCTTTTTACAGTT
扩增引物 Amplification primers	CRISPR-SlPL-F	ACGAAGGAACAAGGGACCCA
扩增引物 Amplification primers	CRISPR-SlPL-R	CGGGTGGAGAACTGTTTCGG
测序引物 Sequencing primers	CRISPR-SlPL-F	CAGATAGGAACAAGGAGT
测序引物 Sequencing primers	CRISPR-SlPL-R	GGTGGAAATGCTATGGTG
	35S-F	TCCCACTGAATCAAAGGC
qPCR引物 Primer for qPCR	Actin-F	CTCTACATACTTGAGAGGTGCC
qPCR引物 Primer for qPCR	Actin-R	AGACGAGGAGAAAACATCACAA
	SlPL-F	GGAAATCCAATCGACGATTGTT
	SlPL-R	CGTTTTTCCCAAATCCAATTGC
	SlCel2-F	TATCAAATGGCGTAGAGACTCC
	SlCel2-R	ACATTATCTCCGGCATCGTAAT
	SlPG2-F	CCAAAGGAATAGTATTCTCCTTCTC
	SlPG2-R	GTTTTTCCATCACCCTTAGCTC
	SlPME2.1-F	ATGCTACCATCATTACTGGGAG
	SlPME2.1-R	GATAAGCATCGATACGACAACG
	SlGH9C2-F	CCAGAAGTACTACCGATCTGTC
	SlGH9C2-R	TCAGTCATAGACCAACCAGTTC
	SlACO1-F	CATACAGACGCAGGAGGCATCA
	SlACO1-R	TAGAGTGGCGCATGGGAGGAA
	SlACS4-F	CTCCTCAAATGGGGAGTACG
	SlACS4-R	TTTTGTTTGCTCGCACTACG
	SlERF2-F	CGACCTATGGCCGACTGATT
	SlERF2-R	GGTCACGAATTTCAGCAGCC

用途 Purpose	基础培养基 Culture medium	蔗糖/ （g · L^-1） Sucrose	琼脂粉/ （g · L^-1） Agaric	反式玉米素/ （mg · L^-1） trans-ZT	IAA/ （mg · L^-1）	IBA /（mg · L^-1）	AS / （mg · L^-1）	特美汀/ （mg · L^-1）Trimethoprim	卡那霉素/ （mg · L^-1）Kanamycin
播种 Seeding	1/2MS	30	7.5	0	0	0	0	0	0
预培养 Pre-culture	MS	30	9	2	0.1	0	0	0	0
悬浮培养 Slave culture	MS	30	0	0	0	0	100	0	0
共培养 Co-culture	MS	30	9	2	0.1	0	100	0	0
筛选培养1 Screen 1	MS	30	9	2	0.1	0	0	200	50
筛选培养2 Screen 2	MS	30	9	0.2	0.1	0	0	400	50
生根 Rooting	1/2MS	30	7.5	0	0	2	0	0	50

用途 Purpose	基础培养基 Culture medium	蔗糖/ （g · L^-1） Sucrose	琼脂粉/ （g · L^-1） Agaric	反式玉米素/ （mg · L^-1） trans-ZT	IAA/ （mg · L^-1）	IBA /（mg · L^-1）	AS / （mg · L^-1）	特美汀/ （mg · L^-1）Trimethoprim	卡那霉素/ （mg · L^-1）Kanamycin
播种 Seeding	1/2MS	30	7.5	0	0	0	0	0	0
预培养 Pre-culture	MS	30	9	2	0.1	0	0	0	0
悬浮培养 Slave culture	MS	30	0	0	0	0	100	0	0
共培养 Co-culture	MS	30	9	2	0.1	0	100	0	0
筛选培养1 Screen 1	MS	30	9	2	0.1	0	0	200	50
筛选培养2 Screen 2	MS	30	9	0.2	0.1	0	0	400	50
生根 Rooting	1/2MS	30	7.5	0	0	2	0	0	50

Study on the Regulatory Mechanism of SlPL Gene Affecting Tomato Fruit Cracking

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