马铃薯中重力响应调节基因鉴定及功能分析

doi:10.13560/j.cnki.biotech.bull.1985.2025-0003

摘要/Abstract

摘要：

目的解析马铃薯（Solanum tuberosum L.）重力响应的分子机理，为马铃薯的遗传改良和育种提供潜在基因资源。方法通过石蜡切片对马铃薯的匍匐茎和地上茎进行组织学结构观察；通过同源比对鉴定马铃薯中的重力响应调控基因，使用RT-qPCR验证其表达水平，利用转录组测序分析其表达模式；以四倍体马铃薯材料Desiree为背景，构建了重力响应关键基因StLAZY1-1的过表达转基因株系，并对其表型进行了观察统计。结果匍匐茎与地上茎的组织结构存在明显的形态学差异，特别是维管组织的排列和淀粉粒沉积两个方面差别显著。重力响应相关基因PRAF/RLD FAMILY MEMBER（RLD）、SCARECROW（SCR）、SHOOT-ROOT（SHR）、BRX-LIKE4（BRXL4）、PIN-FORMED3（PIN3）在不同物种中具有保守的结构域和分布特征，LAZY家族的结构域分布则具有多样性。重力响应相关基因的表达具有组织特异性，并且表达水平受到光信号调控。通过遗传转化验证马铃薯StLAZY1-1基因的功能，发现与野生型相比StLAZY1-1过表达株系在株高、匍匐茎数量上都具有显著变化。结论马铃薯中关键重力响应调节基因StLAZY1-1影响马铃薯的株高和匍匐茎数量。

关键词: 马铃薯, 重力响应, LAZY1基因, 石蜡切片

Abstract:

Objective This study aims to provide a reference for elucidating the molecular mechanisms underlying gravity response in potato (Solanum tuberosum L.) and to offer potential genetic resources for the genetic improvement and breeding of potato. Method Paraffin sectioning was employed to examine the histological structures of potato stolons and aerial stems. Gravity-responsive regulatory genes in potato were identified through homology alignment, with their expressions validated by RT-qPCR. Expression patterns were further analyzed using transcriptome sequencing. Utilizing the tetraploid potato variety Desiree as a background, we constructed transgenic lines that overexpressed the gravity-responsive key gene StLAZY1-1, followed by observation and statistical analysis of their phenotypes. Result Significant morphological differences in tissue structure were observed between the stolon and aerial stem, particularly regarding the arrangement of vascular tissues and the deposition of starch granules. Gravity-responsive genes, including PRAF/RLD FAMILY MEMBER (RLD), SCARECROW (SCR), SHOOT-ROOT (SHR), BRX-LIKE4 (BRXL4), and PIN-FORMED3 (PIN3), demonstrated conserved domains and distribution patterns across various species, whereas LAZY family genes displayed diverse domain distributions. The expressions of gravity-responsive genes were found to be tissue-specific and regulated by light signals. Functional verification of the StLAZY1-1 gene in transgenic potato lines demonstrated significant alterations in plant height and stolon number when compared to wild-type plants. Conclusion The key gravity-responsive regulatory gene StLAZY1-1 in potato influences plant height and the number of stolons.

Key words: potato, gravity response, LAZY1-1 gene, paraffin sectioning

罗稷林, 栗锦烨, 贾玉鑫. 马铃薯中重力响应调节基因鉴定及功能分析[J]. 生物技术通报, 2025, 41(6): 109-118.

LUO Ji-lin, LI Jin-ye, JIA Yu-xin. Identification and Functional Analysis of Gravity Response Regulatory Genes in Potato[J]. Biotechnology Bulletin, 2025, 41(6): 109-118.

图/表 7

表 1 RT-qPCR 引物序列

Table 1 Primer sequences for RT-qPCR

基因名称Gene name	引物名称 Primer name	引物序列Primer sequence（5'-3'）
StACTIN	Stactin-F	GGGATGGAGAAGTTTGGTGGTGG
StACTIN	Stactin-R	CTTCGACCAAGGGATGGTGTAGC
StBRXL4	BRXL4-qRT-F	TGCCAGTGATGTTGAGACTGA
StBRXL4	BRXL4-qRT-R	TGGGCAGTGCAGGATCTTTT
StLAZY1-1	LAZY1-1-qRT-F	TTCAGCAGAGCCAAAACCCC
StLAZY1-1	LAZY1-1-qRT-R	TTTGAGAGGCGATGGTGAGG
StLAZY2	LAZY2-qRT-F	GCGAGACACACTTCCCGAAT
StLAZY2	LAZY2-qRT-R	ATCCGAATCGGTTTTGACCCA
StLAZY5	LAZY5-qRT-F	CGGTGGAGAAAAGAAAGAACGA
StLAZY5	LAZY5-qRT-R	TTGCCCTGTTCCTGAAGTTGA
StLAZY6	LAZY6-qRT-F	GCCATAATAGCACCAGCAGTTT
StLAZY6	LAZY6-qRT-R	TGGCGTTGGAGGGTCAGTA
StPIN3	PIN3-qRT-F	TCGGATGTTGGTTTCTGACCA
StPIN3	PIN3-qRT-R	ATGTGGAGCTAGACCCCAGT
StRLD1	RLD1-qRT-F	TTCTCTGCCTTCTGGCAACC
StRLD1	RLD1-qRT-R	CTTGGTGCTCCCCGAATCTT
StRLD2	RLD2-qRT-F	GCTGGAGAAGAGACTGCCAA
StRLD2	RLD2-qRT-R	GCAGTGAGATTGGAACCCGA
StSCR	SCR-qRT-F	GCCGAGAGATTAGGCCTTCC
StSCR	SCR-qRT-R	ACCTTCGGAGCCAACCTTTG
StSHR	SHR-qRT-F	GACACAACAATCTCAATCTCATCA
StSHR	SHR-qRT-R	ACATCCACATGAGTTGTTGGAC

图1 马铃薯地上茎和匍匐茎的组织切片A：幼嫩茎；B：成熟茎；C：匍匐茎

Fig. 1 Anatomy of potato stem and stolonA: Young stem. B: Mature stem. C: Creeping stem

图2 重力响应调控基因结构域比对

Fig. 2 Aligment of domains in gravity response regulatory genes

图3 马铃薯中重力响应调控基因表达模式图A：RT-qPCR检测马铃薯各组织表达模式；B：RNA-seq检测马铃薯各组织表达模式。LD代表长日照，SD代表短日照

Fig. 3 Expression pattern of gravity-responsive regulatory genes in potatoA: RT-qPCR detects expression pattern in tissues of potato; B: RNA-seq detects expression pattern in tissues of potato. LD refers to long day, SD refers to short day

图4 LAZY家族蛋白进化树At: Arabidopsis thaliana; Sl: Solanum lycopersicum; St: Solanum tuberosum; Os: Oryza sativa; Zm: Zea mays

Fig. 4 Phylogenetic tree of proteins in the LAZY family

图5 马铃薯不同发育形态的匍匐茎中StLAZY1-1表达模式A：马铃薯部分地下组织示意图；B：RT-qPCR检测StLAZY1-1各组织表达水平，使用单因素方差检验，*P<0.05，**P<0.01，***P<0.001，****P<0.000 1，下同

Fig. 5 StLAZY1-1 expression patterns in the stolon of different developmental morphology of potatoA: Schematic diagram of underground organization of potato part. B: The expression of StLAZY1-1 was detected by RT-qPCR, one-way ANOVA was used, *P<0.05, **P<0.01,***P<0.001, ****P<0.000 1, the same below

图6 LAZY1-1OE阳性植株鉴定与表型统计A：Desiree背景LAZY1-1阳性苗检测；B：Desiree 背景转基因苗表型统计

Fig. 6 Identification and phenotypic statistics of LAZY1-1OE positive plantsA: Detection of LAZY1-1-positive plants in the Desiree background; B: Phenotypic statistics of transgenic plants in the Desiree background

参考文献 32

33	Xu R, Sun CQ. What happened during domestication of wild to cultivated rice [J]. Crop J, 2021, 9(3): 564-576.
34	Li PJ, Wang YH, Qian Q, et al. LAZY1 controls rice shoot gravitropism through regulating polar auxin transport [J]. Cell Res, 2007, 17(5): 402-410.
35	Yoshihara T, Spalding EP. LAZY genes mediate the effects of gravity on auxin gradients and plant architecture [J]. Plant Physiol, 2017, 175(2): 959-969.
1	Vandenbrink JP, Kiss JZ. Plant responses to gravity [J]. Semin Cell Dev Biol, 2019, 92: 122-125.
2	Volkmann D, Baluska F. Gravity: one of the driving forces for evolution [J]. Protoplasma, 2006, 229(2-4): 143-148.
3	Hangarter RP. Gravity, light and plant form [J]. Plant Cell Environ, 1997, 20(6): 796-800.
4	ARWIN F. The statolith-theory of geotropism [J]. Proc R Soc Lond, 1903, 71(467-476): 362-373.
5	Roychoudhry S, Kepinski S. Shoot and root branch growth angle control-the wonderfulness of lateralness [J]. Curr Opin Plant Biol, 2015, 23: 124-131.
6	Emerson R, Beadle G, Fraser AC. A summary of linkage studies in maize [J]. Cornell Agricultural Experiment, 1935,180:83.
7	Dong ZB, Jiang C, Chen XY, et al. Maize LAZY1 mediates shoot gravitropism and inflorescence development through regulating auxin transport, auxin signaling, and light response [J]. Plant Physiol, 2013, 163(3): 1306-1322.
8	Fukaki H, Fujisawa H, Tasaka M. SGR1, SGR2, SGR3: novel genetic loci involved in shoot gravitropism in Arabidopsis thaliana [J]. Plant Physiol, 1996, 110(3): 945-955.
9	Zhang H, Li X, Sang DJ, et al. PROG1 acts upstream of LAZY1 to regulate rice tiller angle as a repressor [J]. Crop J, 2023, 11(2): 386-393.
10	Chen JY, Yu RB, Li N, et al. Amyloplast sedimentation repolarizes LAZYs to achieve gravity sensing in plants [J]. Cell, 2023, 186(22): 4788-4802.e15.
11	Hollender CA, Hill JL Jr, Waite J, et al. Opposing influences of TAC1 and LAZY1 on lateral shoot orientation in Arabidopsis [J]. Sci Rep, 2020, 10(1): 6051.
12	Taniguchi M, Furutani M, Nishimura T, et al. The Arabidopsis LAZY1 family plays a key role in gravity signaling within statocytes and in branch angle control of roots and shoots [J]. Plant Cell, 2017, 29(8): 1984-1999.
13	Yoshihara T, Spalding EP, Iino M. AtLAZY1 is a signaling component required for gravitropism of the Arabidopsis thaliana inflorescence [J]. Plant J, 2013, 74(2): 267-279.
14	Furutani M, Hirano Y, Nishimura T, et al. Polar recruitment of RLD by LAZY1-like protein during gravity signaling in root branch angle control [J]. Nat Commun, 2020, 11(1): 76.
15	Che XM, Splitt BL, Eckholm MT, et al. BRXL4-LAZY1 interaction at the plasma membrane controls Arabidopsis branch angle and gravitropism [J]. Plant J, 2023, 113(2): 211-224.
16	Caspar T, Pickard BG. Gravitropism in a starchless mutant of Arabidopsis: implications for the starch-statolith theory of gravity sensing [J]. Planta, 1989, 177: 185-197.
17	Vitha S, Yang M, Sack FD, et al. Gravitropism in the starch excess mutant of Arabidopsis thaliana [J]. Am J Bot, 2007, 94(4): 590-598.
18	Zhu L, Yin T, Zhang MJ, et al. Genome-wide identification and expression pattern analysis of the kiwifruit GRAS transcription factor family in response to salt stress [J]. BMC Genomics, 2024, 25(1): 12.
19	Fukaki H, Wysocka-Diller J, Kato T, et al. Genetic evidence that the endodermis is essential for shoot gravitropism in Arabidopsis thaliana [J]. Plant J, 1998, 14(4): 425-430.
20	Johri MM. Hormonal regulation in green plant lineage families [J]. Physiol Mol Biol Plants, 2008, 14(1-2): 23-38.
21	Friml J, Wiśniewska J, Benková E, et al. Lateral relocation of auxin efflux regulator PIN3 mediates tropism in Arabidopsis [J]. Nature, 2002, 415(6873): 806-809.
22	Zažímalová E, Křeček P, Skůpa P, et al. Polar transport of the plant hormone auxin-the role of PIN-FORMED (PIN) proteins [J]. Cell Mol Life Sci, 2007, 64(13): 1621-1637.
23	Horton D. Underground crops: Long-term trends in production of roots and tubers [J]. Winrock International, 1988,7-9.
24	Navarro C, Abelenda JA, Cruz-Oró E, et al. Control of flowering and storage organ formation in potato by FLOWERING LOCUS T [J]. Nature, 2011, 478(7367): 119-122.
25	Zierer W, Rüscher D, Sonnewald U, et al. Tuber and tuberous root development [J]. Annu Rev Plant Biol, 2021, 72: 551-580.
26	Xu X, Vreugdenhil D, Lammeren AAMV. Cell division and cell enlargement during potato Tuber formation [J]. J Exp Bot, 1998, 49(320): 573-582.
27	Abelenda JA, Bergonzi S, Oortwijn M, et al. Source-sink regulation is mediated by interaction of an FT homolog with a SWEET protein in potato [J]. Curr Biol, 2019, 29(7): 1178-1186.e6.
28	Tang D, Jia YX, Zhang JZ, et al. Genome evolution and diversity of wild and cultivated potatoes [J]. Nature, 2022, 606(7914): 535-541.
29	叶明旺, 张春芝, 黄三文. 二倍体栽培马铃薯高效遗传转化体系的建立 [J]. 中国农业科学, 2018, 51(17): 3249-3257.
	Ye MW, Zhang CZ, Huang SW. Construction of high efficient genetic transformation system for diploid potatoes [J]. Sci Agric Sin, 2018, 51(17): 3249-3257.
30	Overbeek JV. “Lazy,” an a-geotropic form of maize [J]. J Hered, 1936, 27(3): 93-96.
31	Howard TP 3rd, Hayward AP, Tordillos A, et al. Identification of the maize gravitropism gene lazy plant1 by a transposon-tagging genome resequencing strategy [J]. PLoS One, 2014, 9(1): e87053.
32	Huang X, Kurata N, Wei X, et al. A map of rice genome variation reveals the origin of cultivated rice[J]. Nature, 2012, 490(7421):497-501.

[1]	宋慧洋, 苏宝杰, 李京昊, 梅超, 宋倩娜, 崔福柱, 冯瑞云. 马铃薯StAS2-15基因的克隆及盐胁迫下功能分析[J]. 生物技术通报, 2025, 41(5): 119-128.
[2]	文博霖, 万敏, 胡建军, 王克秀, 景晟林, 王心悦, 朱博, 唐铭霞, 李兵, 何卫, 曾子贤. 马铃薯川芋50遗传转化及基因编辑体系的建立[J]. 生物技术通报, 2025, 41(4): 88-97.
[3]	刘涛, 王志淇, 吴文博, 石文婷, 王超楠, 杜崇, 杨中敏. 马铃薯GRAM基因家族鉴定与表达分析[J]. 生物技术通报, 2025, 41(4): 145-155.
[4]	张益瑄, 马宇, 王童童, 盛苏奥, 宋家凤, 吕钊彦, 朱晓彪, 侯华兰. 马铃薯DIR家族全基因组鉴定及表达模式分析[J]. 生物技术通报, 2025, 41(3): 123-136.
[5]	俞婷, 黄丹丹, 朱炎辉, 杨梅宏, 艾菊, 高冬丽. 马铃薯Stpatatin 05基因转录调控因子筛选及互作验证[J]. 生物技术通报, 2025, 41(3): 137-145.
[6]	覃悦, 杨妍, 张磊, 卢丽丽, 李先平, 蒋伟. 二倍体和四倍体马铃薯StGAox基因鉴定与比较分析[J]. 生物技术通报, 2025, 41(3): 146-160.
[7]	申鹏, 高雅彬, 丁红. 马铃薯SAT基因家族的鉴定和表达分析[J]. 生物技术通报, 2024, 40(9): 64-73.
[8]	宋兵芳, 柳宁, 程新艳, 徐晓斌, 田文茂, 高悦, 毕阳, 王毅. 马铃薯G6PDH基因家族鉴定及其在损伤块茎的表达分析[J]. 生物技术通报, 2024, 40(9): 104-112.
[9]	王超, 白如仟, 管俊梅, 罗稷林, 何雪姣, 迟绍轶, 马玲. 马铃薯块茎变绿中StHY5对龙葵素合成的促进作用[J]. 生物技术通报, 2024, 40(9): 113-122.
[10]	夏士轩, 耿泽栋, 祝光涛, 张春芝, 李大伟. 基于深度学习的马铃薯花粉活力快速检测[J]. 生物技术通报, 2024, 40(9): 123-130.
[11]	毛向红, 卢瑶, 范向斌, 杜培兵, 白小东. 基于SSR荧光标记毛细管电泳的马铃薯品种遗传多样性分析及分子身份证构建[J]. 生物技术通报, 2024, 40(9): 131-140.
[12]	袁兰, 黄娅楠, 张贝妮, 熊雨萌, 王洪洋. 基于流式细胞仪鉴定马铃薯倍性的高通量样品制备方法[J]. 生物技术通报, 2024, 40(9): 141-147.
[13]	宋倩娜, 段永红, 冯瑞云. CRISPR/Cas9介导的高效四倍体马铃薯试管薯基因编辑体系的建立[J]. 生物技术通报, 2024, 40(9): 33-41.
[14]	王柯然, 闫俊杰, 刘建凤, 高玉林. RNAi技术在马铃薯害虫防控中的应用和风险[J]. 生物技术通报, 2024, 40(9): 4-10.
[15]	张小妹, 周南伶, 张赛行, 王超, 沈玉龙, 管俊梅, 马玲. 马铃薯StDREBs基因的克隆及其表达分析[J]. 生物技术通报, 2024, 40(9): 42-50.