矮牵牛花朵大小遗传规律及相关基因的表达分析

doi:10.11983/CBB23141

摘要/Abstract

摘要： 花朵大小是植物进化及物种形成过程中的关键因子, 也是决定植物观赏价值的重要性状, 研究其遗传规律与调控机制具有重要的科学意义和实用价值。以不同花朵大小的矮牵牛(Petunia hybrida)自交系和原生种为材料, 配制大花×中花和大花×小花杂交组合, 构建遗传群体, 研究其花朵大小的遗传规律。结果表明, 大花自交系W与腋花矮牵牛S26 (中花)杂交F₁群体均为大花, F₂群体中大花与中花的分离比接近3:1, BC₁回交群体中大花与中花的分离比接近1:1; 大花自交系W与中花自交系S杂交F₁群体均为大花, F₂群体花朵大小出现分离, 大花与中花的比例接近2:1; 大花自交系W与膨大矮牵牛S6 (小花)杂交F₁群体均为中花, F₂群体则出现花径从大到小的连续分布。利用主基因+多基因混合模型分析, 按照AIC值最小的标准选出W × S26组合的最优模型为1MG-AD, W × S的最优模型为2MG-EAD, 由此判定W大花对S26中花由1对主显性基因控制, 具加性-显性效应, W大花对S中花则由2对主基因控制, 具等加性-显性效应。此外, 根据前期大、小花转录组分析结果, 选取9个可能调控花朵大小的基因, 通过qPCR检测了它们在不同花朵大小的矮牵牛株系花瓣中的表达水平, 结果发现细胞分裂素受体基因PhHK以及响应细胞分裂素信号的type-A RRs基因在大花中的表达水平普遍高于中花和小花, 表明细胞分裂素信号途径可能是参与调控矮牵牛大花性状的关键因素。

关键词: 矮牵牛, 花朵大小, 遗传规律, 细胞分裂素信号途径

Abstract: Flower size is a key factor in plant evolution and speciation, and also an important trait that determines plant ornamental value, so it is of great scientific significance and practical value to study the inheritance law and regulatory mechanism of floral size. To clarify the inheritance law of flower size in petunia, the inbred lines and wild species of Petunia hybrida with different flower sizes were used to make cross combinations and construct genetic populations in this study, including large-flowered line × medium-flowered lines (W × S26 and W × S) and large-flowered line × small-flowered line (W × S6). The results showed that all F₁ generation of W × S26 were large-flowered plants, while the flower size appeared separation in F₂ population with the ratio between large-and medium-flowered individuals of about 3:1, and the segregation ratio between large- and medium-flowered plants was close to 1:1 in the BC₁ backcross population. For the W × S combination, all F₁ individuals were large-flowered, while the flower size appeared separation in the F₂ population, with large- to medium-flowered plants close to 2:1. The F₁ progenies of W × S6 are all medium-flowered plants, while the flower size of the F₂ generation showed evident variation and continuous distribution. Performing mixed major gene plus polygene inheritance model analysis, the optimal models for W × S26 and W × S combination were 1MG-AD and 2MG-EAD, respectively, according to the standard of minimum AIC value. It is reasonable to conclude that the large flower trait of the inbred line W is controlled by a single dominant gene related to the middle flower trait of S26, with additive dominant effect, whereas the large flower of the inbred line W is controlled by two major genes related to the small flower of the inbred lines with equal additive dominant effect. In addition, nine genes that may regulate flower size of petunia were selected based on the transcriptomic analysis of large and small flowers, and their expression levels were detected in the petals of various strains with different flower sizes by qRT-PCR. The results showed that the expression levels of cytokinin receptor gene PhHK and Type-A RRs in response to cytokinin signal were generally higher in large flowers than in medium and small flowers, suggesting that cytokinin signaling pathway may be a key factor involved in regulating the large flower trait in petunia.

Key words: petunia, flower size, inheritance law, cytokinin signaling pathway

孙苗苗, 张蔚, 张林霞, 霍竣涛, 李志能, 刘国锋. 矮牵牛花朵大小遗传规律及相关基因的表达分析. 植物学报, 2024, 59(3): 422-432.

Miaomiao Sun, Wei Zhang, Linxia Zhang, Juntao Huo, Zhineng Li, Guofeng Liu. Inheritance Analysis of Flower Size and Expression of Related Genes in Petunia hybrida. Chinese Bulletin of Botany, 2024, 59(3): 422-432.

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链接本文: https://www.chinbullbotany.com/CN/10.11983/CBB23141

https://www.chinbullbotany.com/CN/Y2024/V59/I3/422

图/表 8

图1 不同花朵大小矮牵牛实验材料 W、DHZ、KZ、W115、S、SW、HC和SP: 矮牵牛自交系; S26: 腋花矮牵牛; S6: 膨大矮牵牛。Bar=1 cm

Figure 1 The tested materials of petunia with different flower sizes W, DHZ, KZ, W115, S, SW, HC, and SP: The inbred lines of Petunia hybrida; S26: P. axillaris; S6: P. inflata. Bar=1 cm

表1 实时荧光定量PCR引物

Table 1 Primers used for quantitative real-time PCR

Gene name	Primer name	Primer sequence (5'−3')
PhARR496	qPhARR496-F	GCAACAACAAAGAAGCAGAGCAGTT
PhARR496	qPhARR496-R	GGTGATGATGGTAATGGCTGTGTCA
PhARR828	qPhARR828-F	CATCTTCCTCGCTCTCGCTCTC
PhARR828	qPhARR828-R	TGAGTCGTCGTGTTGGTGAATGT
PhARR970	qPhARR970-F	ACTCCTCTTCCTCATCAGTCATCATCA
PhARR970	qPhARR970-R	GCTGCTGCCATTCCCATTGTTG
PhARR1029	qPhARR1029-F	GCAAGAGCATGAACAGAACAAGTCA
PhARR1029	qPhARR1029-R	CCAGGCATACAGTAGTCCGTGATAA
PhARGOS057	qPhARGOS057-F	TCTCTTCCATCATACGCTTTGGTCAA
PhARGOS057	qPhARGOS057-R	AGGCAAGAACCATAGGAGATTGTTGA
PhARGOS459	qPhARGOS459-F	GCACATGGAATCATCAGCAGAAGC
PhARGOS459	qPhARGOS459-R	GGTGGCAATGGTGGAAGGATCAA
PhHK	qPhHK-F	CCAAGTTCTTTGAATCCAAGCCTCAC
PhHK	qPhHK-R	AATAAGAATGCAGCACCAGCAGAATG
PhTCP3b	qPhTCP3b-F	CCTCTCTGCTCACACAGCCATTC
PhTCP3b	qPTCP3b-R	GCCCAGCAATACTGTCAACTAAACT
PhTCP4a	qPhTCP4a-F	TGACCAAGCAGCCCTTTTCTCT
PhTCP4a	qPhTCP4a-R	TGGAACTCTGTGGTGAAATGTCTGT

图2 W × S26亲本及不同后代群体花径的频数分布 (A) 母本W; (B) 父本S26; (C) F1群体; (D) F2群体; (E) BC1群体; (F) 大花BCF2群体; (G) 中花BCF2群体

Figure 2 Frequency distribution histogram of flower diameter for parents and filial generations of W × S26 (A) Female parent W; (B) Male parent S26; (C) F1 generation; (D) F2 generation; (E) BC1 generation; (F) BCF2 generation from large flower; (G) BCF2 generation from medium flower

图3 W × S和W × S6亲本及F1、F2群体花径频数的分布 (A) 亲本S; (B) 亲本S6; (C) W × S F1群体; (D) W × S F2群体; (E) W × S6 F1群体; (F) W × S6 F2群体

Figure 3 Frequency distribution histogram of flower diameter for parent, F1, and F2 generations of W × S and W × S6 (A) Parent S; (B) Parent S6; (C) F1 generation of W × S; (D) F2 generation of W × S; (E) F1 generation of W × S6; (F) F2 generation of W × S6

表2 各遗传模型极大似然值(MLV)、AIC (Akaike information criterion)值和达到显著水平个数

Table 2 Maximum likelihood value (MLV), Akaike information criterion (AIC) value, and the number of reaching significant levels for each genetic model

Cross combination	Genetic model	Maximum likelihood value	AIC	Number of significant levels (P<0.05)
Cross combination	Genetic model	Maximum likelihood value	AIC	U₁²	U₂²	U₃²	nW²	D_n
W × S26	1MG-AD	-599.9208	1207.8420	0	0	0	0	0
	2MG-ADI	-596.0287	1212.0570	0	0	0	0	0
	MX1-AD-AD	-596.8717	1211.7440	0	0	0	0	0
W × S	2MG-EAD	-242.0814	492.1628	0	0	0	0	0
	MX2-EAD-AD	-245.5385	495.0770	0	0	0	0	0
	2MG-EA	-244.9837	497.9675	0	0	0	0	0

表3 矮牵牛杂交后代群体花朵大小分离卡方检验

Table 3 Chi square test for flower size segregation in the hybrid offsprings of petunia

Cross combination (female × male)	Population types	Plant numbers	Large flower	Medium flower	Small flower	Theoretical ratio	Actual ratio	(χc²) Chi-square test	P
W × S26	F₁	32	32	0	0	-	-	-	-
	F₂	134	98	36	0	3:1	3:1	0.249	0.618
	BC₁	129	67	62	0	1:1	1:1	0.194	0.660
	Large flower BCF₂	259	194	65	0	3:1	3:1	0.001	0.971
	Medium flower BCF₂	44	0	44	0	-	-	-	-
W × S	F₁	72	72	0	0	-	-	-	-
W × S	F₂	115	76	39	0	-	-	-	-
W × S6	F₁	57	0	57	0	-	-	-	-
W × S6	F₂	200	11	168	21	-	-	-	-

图4 W × S26花朵大小遗传规律图解 Bar=1 cm

Figure 4 Diagram of genetic law of flower size for W × S26 Bar=1 cm

图5 参与调控花朵大小的相关基因在不同矮牵牛株系中的表达水平误差线代表3个生物学重复的标准误差。不同小写字母表示不同株系间差异显著。

Figure 5 Expression levels of flower size regulating related genes in different strains of petunia Vertical bars represent SE of three biological replicates. Different lowercase letters indicate significant differences among strains.

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