Identification and Expression Analysis of 13 Gene Families in the Chestnut Flavonoid Synthesis Pathway

doi:10.13560/j.cnki.biotech.bull.1985.2024-0537

Abstract

Abstract:

Objective Identifying 13 gene families of the flavonoid synthesis pathway from the chestnut genome and analyzing their expression patterns during fruit ripening and PCD of fruit top bud tissue may lay a foundation for subsequent studies on the function of flavonoids-related gene in chestnut. Method Bioinformatics techniques was used to identify 13 gene families in chestnut flavonoids synthesis pathway, and analyze their phylogenetic relationships, protein physicochemical properties, protein structure, gene structure, cis-acting elements, collinearity relationships, and codon preference. Both transcriptome and RT-qPCR were used to analyze the expression patterns during fruit ripening and PCD of fruit top bud tissue. Result The 57 members of 13 chestnut flavonoid synthesis pathway gene family distributed on 12 chromosomes and 4 contigs. Each gene family contained 1-2 key domains, and the number of introns of each gene families member was similar. The results of cis-acting element analysis showed that a large number of growth and development related elements, hormone response elements and stress response elements existed on the promoters of flavonoid synthesis pathway gene family members in chestnut. A total of seven collinear gene pairs were found within the chestnut genome, and four genes were also found to have collinear genes with eight monodicotydon plants. Codon usage bias analysis found preferred A/U endings with RSCU values greater than 1, and all ENC values were above 35. Both transcriptome and RT-qPCR results showed the upregulation of multiple flavonoids synthesis pathway genes during chestnut fruit ripening and PCD of fruit top bud tissues. Conclusion A total of 57 genes of the chestnut flavonoid synthesis pathway were identified. The genes of CmLAR2, CmCHI1, Cm4CL1, CmC4H3, CmPAL1, CmPAL2, Cm4CL6 and CmANR3 were upregulated in chestnut fruit ripening process and PCD of fruit top bud tissue, and these genes may be involved in chestnut fruit ripening process and PCD of chestnut bud tissue.

Key words: chestnut, flavonoids, gene family, collinearity, codon

YANG Yong, CAO Rui, KANG Xiao-xiao, LIU Jing, WANG Xuan, ZHANG Hai-e. Identification and Expression Analysis of 13 Gene Families in the Chestnut Flavonoid Synthesis Pathway[J]. Biotechnology Bulletin, 2025, 41(2): 270-283.

Figures/Tables 13

References 42

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基因 Gene	正向引物 Forward primer （5′-3′）	反向引物 Reverse primer （5′-3′）
CmLAR2	GTGGAACCAGCCCTGACC	GGTAGGGCCAAGCAGCAA
CmCHI1	AGCGTCTCTTCCGGGAGT	CGCCGAGGAACAGTGTGT
Cm4CL1	TGACTCAGGCCGACGAGA	CGACTTGTTGTGCCACGC
CmC4H3	AGGGTGTGGAGTTCGGGT	ATGCTCGCCGTAAACCGT
CmPAL1	AGGCCCTCCAATGCAACC	CGGCAGTGACCTGAGCAA
CmPAL2	CGACTCGTGCCGCTATGT	CGGAGTGGCAAGCATGGA
Cm4CL6	ATCCAGAGGCCACGGCTA	CTCGGCAGGGGCTACTTG
CmANR3	CACCGTTCGGTCTGACCC	GGGTGCATTGAAACTCTCTGG
CmActin	ATTCACGAGACCACCTACA	TGCCACAACCTTAATCTTCAT

基因 Gene	正向引物 Forward primer （5′-3′）	反向引物 Reverse primer （5′-3′）
CmLAR2	GTGGAACCAGCCCTGACC	GGTAGGGCCAAGCAGCAA
CmCHI1	AGCGTCTCTTCCGGGAGT	CGCCGAGGAACAGTGTGT
Cm4CL1	TGACTCAGGCCGACGAGA	CGACTTGTTGTGCCACGC
CmC4H3	AGGGTGTGGAGTTCGGGT	ATGCTCGCCGTAAACCGT
CmPAL1	AGGCCCTCCAATGCAACC	CGGCAGTGACCTGAGCAA
CmPAL2	CGACTCGTGCCGCTATGT	CGGAGTGGCAAGCATGGA
Cm4CL6	ATCCAGAGGCCACGGCTA	CTCGGCAGGGGCTACTTG
CmANR3	CACCGTTCGGTCTGACCC	GGGTGCATTGAAACTCTCTGG
CmActin	ATTCACGAGACCACCTACA	TGCCACAACCTTAATCTTCAT

基因家族 Gene family	以马可夫模型编号 Pfam ID	结构域名称 Domain name
PAL	PF00221	芳香族氨基酸裂解酶 Aromatic amino acid lyase
4CL	PF13193， PF00501	AMP结合酶C末端结构域，AMP结合酶结构域 AMP-binding enzyme C-terminal domain， AMP-binding enzyme domain
CHS	PF02797， PF00195	查尔酮和二苯乙烯合酶，N/C末端结构域 Chalcone and stilbene synthases， N/C-terminal domain
CHI	PF02431	查尔酮黄烷酮异构酶 Chalcone-flavanone isomerase
LAR	PF05368	NmrA-like结构域 NmrA-like domain
DFR	PF01370	NAD依赖性差向异构酶结构域 NAD dependent epimerase domain
ANR	PF01370	NAD依赖性差向异构酶结构域 NAD dependent epimerase domain
F3H	PF03171， PF14226	2OG-FeII_Oxy结构域，DIOX_N结构域 2OG-FeII_Oxy domain， DIOX_N domain
FLS
ANS
C4H	PF00067	细胞色素P450 Cytochrome P450
F3′5′H
FNSII

基因家族 Gene family	以马可夫模型编号 Pfam ID	结构域名称 Domain name
PAL	PF00221	芳香族氨基酸裂解酶 Aromatic amino acid lyase
4CL	PF13193， PF00501	AMP结合酶C末端结构域，AMP结合酶结构域 AMP-binding enzyme C-terminal domain， AMP-binding enzyme domain
CHS	PF02797， PF00195	查尔酮和二苯乙烯合酶，N/C末端结构域 Chalcone and stilbene synthases， N/C-terminal domain
CHI	PF02431	查尔酮黄烷酮异构酶 Chalcone-flavanone isomerase
LAR	PF05368	NmrA-like结构域 NmrA-like domain
DFR	PF01370	NAD依赖性差向异构酶结构域 NAD dependent epimerase domain
ANR	PF01370	NAD依赖性差向异构酶结构域 NAD dependent epimerase domain
F3H	PF03171， PF14226	2OG-FeII_Oxy结构域，DIOX_N结构域 2OG-FeII_Oxy domain， DIOX_N domain
FLS
ANS
C4H	PF00067	细胞色素P450 Cytochrome P450
F3′5′H
FNSII

复制类型 Duplication types	基因对 Gene pairs		非同义替换率 Ka	同义替换率 Ks	非同义替换率/同义替换率 Ka/Ks
WGD	CmPAL3	CmPAL2	0.123 822 763	1.465 415 048	0.084 496 719
WGD	Cm4CL2	Cm4CL11	0.383 354 993	1.384 646 007	0.276 861 372
WGD	CmANR5	CmANR2	0.549 975 334	2.288 010 781	0.240 372 702
WGD	CmC4H1	CmC4H3	0.085 827 583	1.945 979 279	0.044 105 086
WGD	CmANS3	CmANS1	0.219 052 838	1.816 044 933	0.120 620 825
WGD	Cm4CL8	Cm4CL2	0.366 520 820	1.302 665 198	0.281 362 257
WGD	Cm4CL8	Cm4CL11	0.372 236 766	1.132 735 123	0.328 617 661
PD	Cm4CL14	Cm4CL15	0.045 708 825	0.165 863 036	0.275 581 749
TD	Cm4CL3	Cm4CL4	0.143 933 551	0.443 981 508	0.324 188 167
TD	CmPAL5	CmPAL6	0.054 029 598	0.093 294 174	0.579 131 530
TD	CmFNSII2	CmFNSII3	0.139 352 161	0.427 416 010	0.326 034 023
TD	CmDFR1	CmDFR2	0.189 572 148	0.623 062 431	0.304 258 672
TD	CmANR3	CmANR4	0.137 094 524	0.393 608 899	0.348 301 384
TD	Cm4CL4	Cm4CL5	0.133 265 871	0.425 828 776	0.312 956 470
TD	Cm4CL9	Cm4CL10	0.103 441 680	0.273 134 833	0.378 720 204
TD	CmPAL4	CmPAL5	0.052 152 683	0.128 965 485	0.404 392 564
TRD	CmCHS1	CmCHS2	0.644 246 280	NaN （high sequence divergence）
TRD	CmCHS3	CmCHS4	0.587 247 232	2.429 515 767	0.241 713 694
TRD	CmLAR2	CmLAR1	0.263 851 247	1.291 720 422	0.204 263 432
TRD	CmF3'5'H1	CmF3'5'H2	0.090 142 821	0.211 897 324	0.425 408 022
TRD	CmFLS2	CmFLS3	0.190 856 593	1.395 097 547	0.136 805 196
TRD	CmANS2	CmANS1	0.636 070 470	3.657 827 926	0.173 892 945
TRD	CmFLS1	CmFLS3	0.467 506 223	3.335 289 988	0.140 169 588
TRD	CmANR7	CmANR6	0.015 019 275	0.051 683 849	0.290 599 013
TRD	Cm4CL6	Cm4CL1	0.148 018 360	1.985 685 394	0.074 542 705