毛竹不同类型愈伤组织比较分析

doi:10.12302/j.issn.1000-2006.202203062

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南京林业大学学报（自然科学版） ›› 2023, Vol. 47 ›› Issue (6): 141-149.doi: 10.12302/j.issn.1000-2006.202203062

毛竹不同类型愈伤组织比较分析

黄碧芸¹^,²(), 卓仁英¹, 乔桂荣¹^,^*()

1.中国林业科学研究院亚热带林业研究所,浙江杭州 311400
2.南京林业大学林草学院,江苏南京 210037

收稿日期:2022-03-28 修回日期:2022-06-19 出版日期:2023-11-30 发布日期:2023-11-23
通讯作者: *乔桂荣(gr_q1982@163.com),副研究员。
基金资助:
中央级公益性科研院所基本科研业务费专项资金项目(CAFYBB2020ZB004)

Comparative analysis of different types of callus in moso bamboo

HUANG Biyun¹^,²(), ZHUO Renying¹, QIAO Guirong¹^,^*()

1. Research Institute of Subtropical Forestry, Chinese Academy of Forestry,Hangzhou 311400, China
2. College of Forestry and Grassland,Nanjing Forestry University, Nanjing 210037,China

Received:2022-03-28 Revised:2022-06-19 Online:2023-11-30 Published:2023-11-23

摘要/Abstract

摘要：

【目的】比较不同类型毛竹(Phyllostachys edulis)愈伤组织在组成细胞形态、细胞周期时相分布比例、基因表达等方面的差异,研究不同类型毛竹愈伤组织的增殖与分化潜力,为毛竹离体再生挑选合适的愈伤组织提供依据。【方法】以毛竹未成熟胚为外植体诱导愈伤,依据外观、质地挑选出白色半透明型、白色致密型和黄色致密型等3种不同类型的毛竹愈伤组织;通过石蜡切片组织细胞学观察、流式细胞仪检测细胞周期时相分布比例,探究3种类型毛竹愈伤组织的增殖与再生能力;利用实时荧光定量PCR (qRT-PCR)对毛竹不同类型愈伤组织中胚性维持及生长调节相关基因的表达量进行分析,筛选可用于后续提高再生效率的候选基因。【结果】第2类白色致密型的毛竹愈伤组织表面及内部具有大量胚性细胞组成的分生组织细胞团,处于DNA合成期(S期)、DNA合成后期(G2期)的细胞比例显著高于其他两类愈伤组织,分别达到细胞总数的8.78%和8.95%;2个PeWUS基因在胚性较强的第2类和第3类愈伤中表达量较高,3个PeGRF基因在第2类愈伤组织中表达量最高。【结论】不同类型毛竹愈伤组织增殖和再生能力存在差异,第2类白色致密的愈伤组织增殖和再生能力更强,更适合作为组织培养材料,筛选出的差异基因可作为提高毛竹再生效率的候选基因。

关键词: 毛竹, 愈伤组织, 组织细胞学, 细胞增殖, 基因表达

Abstract:

【Objective】 The proliferation ability and differentiation potential of different types of moso bamboo callus were compared with reference to three aspects: cell morphology, cell cycle phase distribution ratio and gene expression. The results provided a basis for selecting suitable callus for in vitro regeneration of moso bamboo. 【Method】 Three different types of callus were selected according to appearance and texture by using the immature embryo of moso bamboo as an explant to induce callus. The proliferation and regeneration ability of three types of moso bamboo callus were investigated by paraffin section and flow cytometry. Real-time PCR (qRT-PCR) was used to analyze the expression of genes related to embryonic maintenance and growth regulation in different callus types, and to select candidate genes that could be used to improve regeneration efficiency in the future. 【Result】 The type Ⅱ of white and dense callus had a large number of embryogenic meristem cells on the surface and inside. The proportion of cells in stage S and G2 in this callus type was significantly higher than that in the other types of callus, reaching 8.78% and 8.95% of the total number of cells, respectively. At the same time, the expression of two PeWUS was higher in type Ⅱ and type Ⅱ callus with strong embryogenesis, and the expression of seven PeGRF genes was the highest in type Ⅱ callus. 【Conclusion】 There were differences in proliferation and regeneration ability of different types of moso bamboo callus. The type Ⅱ of white dense callus with stronger proliferation and regeneration ability was more suitable as a tissue culture material. The selected differential genes could be used as candidate genes to improve regeneration efficiency of moso bamboo.

Key words: moso bamboo, callus, histology, cell proliferation, gene expression

中图分类号:

S718

黄碧芸,卓仁英,乔桂荣. 毛竹不同类型愈伤组织比较分析[J]. 南京林业大学学报（自然科学版）, 2023, 47(6): 141-149.

HUANG Biyun, ZHUO Renying, QIAO Guirong. Comparative analysis of different types of callus in moso bamboo[J].Journal of Nanjing Forestry University (Natural Science Edition), 2023, 47(6): 141-149.DOI: 10.12302/j.issn.1000-2006.202203062.

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基因 gene	基因ID gene ID	正向引物(5'-3') forward primer(5'-3')	反向引物(5'-3') reverse primer(5'-3')
	PH02Gene04456.t1	TGGAGTGGGACAATGCCAAG	ATGGAGATCTGCTTGCGGAG
	PH02Gene03707.t1	TTTTACTGCGCCAAGAACGC	TCGTCCGTCATCACCTTGAC
WUS	PH02Gene44184.t1	GCGACCATCACGGTGTTCAT	CATACTCGTTGACGGGGAGG
	PH02Gene15929.t1	GGCAGATGGGGTTTGTGGAA	CGAACATGGCTCGCAAGTCT
	PH02Gene32607.t1	AGGCACTCCACTGCACCTGT	CCAGAATGGACACCGAAGAA
	PH02Gene02258.t2	TGGGTACGGGTCCTACTTCG	TGCGTTTCCACAGGCTTTCT
	PH02Gene06385.t2	ATGACCAGGCTTACATTGCG	GTCCGTCGTCAAGTGTTTCC
	PH02Gene10190.t3	GAGCTACCCTCTGCTTGGAA	AGGAGTGGCTGTTGTCATCG
	PH02Gene23215.t2	CGTCGCAGATTCATGTCGAT	GTGATGGCTCAAGGTGGGA
	PH02Gene26399.t1	CCTCGTTTCATCGGATTCAC	CCTGGGCCAGTCCACATAGT
	PH02Gene29026.t2	AAACACTAGGGGCTTCACCA	TGACGGCTCAAGGTAGGAAT
GRF	PH02Gene34199.t1	CACGCCTTCTTCTCCGACAG	TTCGTCGTCCTCTTCTTTGG
	PH02Gene01484.t1	GCTCCCCCGCCTCTCCTGT	CGGCGAAGTAGCGGTAGATG
	PH02Gene03343.t1	ACTCGCCGCAGCCACCGT	CAAGCCCAGCCCAAGAACCAG
	PH02Gene03768.t1	GTCAACTCGCTTCCAAAGGTCG	TGGCAACGGAGAAGTCAGAGG
	PH02Gene06203.t1	CGGACGGACGGCAAGAAATG	TGCTGCTGATGGTTGATGGTGAG
	PH02Gene08777.t1	TGTGAAGCAGGAGAACCAGACGC	GAAGTCAGAGGACGCCATCGGTA
	PH02Gene33803.t1	CATCAAATGAAGGAAACAGTGCG	AAGGCGAGGGGCAACTTTTATT
	PH02Gene45969.t1	CCGCAGCATCAGCAGCACTT	GATGGGTAGGCACAGGAACACG
内参reference gene	PeTIP41	AAAATCATTGTAGGCCATTGTCG	ACTAAATTAAGCCAGCGGGAGTG

毛竹不同类型愈伤组织比较分析

Comparative analysis of different types of callus in moso bamboo

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