植物寡核苷酸荧光原位杂交技术方法

doi:10.11983/CBB22265

植物学报 ›› 2023, Vol. 58 ›› Issue (2): 274-284.DOI: 10.11983/CBB22265

植物寡核苷酸荧光原位杂交技术方法

张凡凡¹, 邢新滢¹, 石文清², 沈懿², 程祝宽²^,³^,^*()

¹北京师范大学生命科学学院, 细胞增殖与调控生物学教育部重点实验室, 北京 100875
²中国科学院遗传与发育生物学研究所, 植物基因组学国家重点实验室及植物基因研究中心, 北京 100101
³扬州大学农学院, 江苏省作物基因组学与分子育种重点实验室/植物功能基因组学教育部重点实验室, 江苏省粮食作物现代生产技术协同创新中心, 扬州 225009

收稿日期:2022-11-20 接受日期:2023-01-10 出版日期:2023-03-01 发布日期:2023-03-15
通讯作者: ^*E-mail: zkcheng@genetics.ac.cn
基金资助:
北京市自然科学基金(6214043);国家自然科学基金(32001522);国家自然科学基金(32000368);中央高校基本科研业务费专项资金(2021NTST20)

Protocals for Oligonucleotide Fluorescence in situ Hybridization in Plants

Fanfan Zhang¹, Xinying Xing¹, Wenqing Shi², Yi Shen², Zhukuan Cheng²^,³^,^*()

¹Key Laboratory of Cell Proliferation and Regulation Biology of Ministry of Education, College of Life Sciences, Beijing Normal University, Beijing 100875, China
²State Key Laboratory of Plant Genomics and Center for Plant Gene Research, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China
³Jiangsu Co-Innovation Center for Modern Production Technology of Grain Crops, Jiangsu Key Laboratory of Crop Genomics and Molecular Breeding/Key Laboratory of Plant Functional Genomics of the Ministry of Education, College of Agriculture, Yangzhou University, Yangzhou 225009, China

Received:2022-11-20 Accepted:2023-01-10 Online:2023-03-01 Published:2023-03-15
Contact: ^*E-mail: zkcheng@genetics.ac.cn

摘要/Abstract

摘要： 寡核苷酸荧光原位杂交技术是一种整合了生物信息学分析、DNA高通量合成以及荧光原位杂交实验的新兴技术。该技术适用于任何具有参考基因组的植物物种, 并可根据研究需求设计靶向某个染色体区域、整条染色体或一组染色体的寡核苷酸探针, 目前已成功应用于数种植物的核型分析、染色体变异、种群演化、同源染色体配对、单倍型分析和异源多倍体识别等研究。该文详述了植物寡核苷酸探针的设计、扩增和标记、染色体制备以及荧光原位杂交的具体操作流程, 以期促进寡核苷酸荧光原位杂交技术在细胞遗传学研究中的应用。

关键词: 荧光原位杂交, 寡核苷酸探针, 染色体, 植物

Abstract: Oligonucleotide fluorescence in situ hybridization (Oligo-FISH) is a new technology that integrates bioinformatics analysis, high throughput DNA synthesis and fluorescence in situ hybridization experiments. It is applicable to plant species with a reference genome. Oligo probes are able design to a chromosomal region, an entire chromosome, and a set of chromosomes according to researchers’ requirements. Oligo-FISH has been successfully applied to studies on karyotyping, chromosome variation, population evolution, homologous chromosome pairing, haplotype analysis, and heteropolyploid identification in several plant species. Here, we introduce the concrete operational processes of oligonucleotide-based probe design, amplification and labeling, chromosomes preparation, and fluorescence in situ hybridization in plants, so as to facilitate Oligo-FISH application in the cytogenetics research in the future.

Key words: fluorescence in situ hybridization, oligonucleotide probe, chromosome, plant

张凡凡, 邢新滢, 石文清, 沈懿, 程祝宽. 植物寡核苷酸荧光原位杂交技术方法. 植物学报, 2023, 58(2): 274-284.

Fanfan Zhang, Xinying Xing, Wenqing Shi, Yi Shen, Zhukuan Cheng. Protocals for Oligonucleotide Fluorescence in situ Hybridization in Plants. Chinese Bulletin of Botany, 2023, 58(2): 274-284.

导出引用管理器 EndNote|Ris|BibTeX

链接本文: https://www.chinbullbotany.com/CN/10.11983/CBB22265

https://www.chinbullbotany.com/CN/Y2023/V58/I2/274

图/表 2

图1 寡核苷酸荧光原位杂交实验流程图 (A) 寡核苷酸探针的扩增和标记; (B) 荧光原位杂交(FISH)实验

Figure 1 Outlines of oligonucleotide fluorescence in situ hybridization (A) Amplification and labeling of oligonucleotide-based probes; (B) Procedures of fluorescence in situ hybridization (FISH)

图2 水稻花粉母细胞减数分裂粗线期染色体的寡核苷酸荧光原位杂交鉴定 (A) 粗线期染色体; (B) 特异性靶向水稻11号染色体短臂的寡核苷酸探针(11 S)信号; (C) 特异性靶向水稻11号染色体长臂的寡核苷酸探针(11 L)信号; (D) 三通道合并图(蓝色为染色体, 绿色为11 S, 红色为11 L)。Bars=5 μm

Figure 2 Characterization of rice meiotic pachytene chromosomes by oligonucleotide fluorescence in situ hybridization (A) Pachytene chromosomes; (B) The signal of oligonucleotide probe specific to the short arm of chromosome 11 in rice (11 S); (C) The signal of oligonucleotide probe specific to the long arm of chromosome 11 in rice (11 L); (D) Three-channel combined panel (Blue: Chromosomes; Green: 11 S; Red: 11 L). Bars=5 μm

参考文献 28

[1]	程新杰, 于恒秀, 程祝宽 (2019). 水稻减数分裂染色体分析方法. 植物学报 54, 503-508. DOI
[2]	Albert PS, Zhang T, Semrau K, Rouillard JM, Kao YH, Wang CJR, Danilova TV, Jiang JM, Birchler JA (2019). Whole-chromosome paints in maize reveal rearrangements, nuclear domains, and chromosomal relationships. Proc Natl Acad Sci USA 116, 1679-1685. DOI PMID
[3]	Beliveau BJ, Joyce EF, Apostolopoulos N, Yilmaz F, Fonseka CY, McCole RB, Chang YM, Li JB, Senaratne TN, Williams BR, Rouillard JM, Wu CT (2012). Versatile design and synthesis platform for visualizing genomes with Oligopaint FISH probes. Proc Natl Acad Sci USA 109, 21301-21306. DOI PMID
[4]	Boyle S, Rodesch MJ, Halvensleben HA, Jeddeloh JA, Bickmore WA (2011). Fluorescence in situ hybridization with high-complexity repeat-free oligonucleotide probes generated by massively parallel synthesis. Chromosome Res 19, 901-909. DOI URL
[5]	Braz GT, do Vale Martins L, Zhang T, Albert PS, Birchler JA, Jiang JM (2020). A universal chromosome identification system for maize and wild Zea species. Chromosome Res 28, 183-194. DOI
[6]	Braz GT, He L, Zhao HN, Zhang T, Semrau K, Rouillard JM, Torres GA, Jiang JM (2018). Comparative Oligo- FISH mapping: an efficient and powerful methodology to reveal karyotypic and chromosomal evolution. Genetics 208, 513-523. DOI URL
[7]	Cheng ZK, Buell CR, Wing RA, Jiang JM (2002). Resolution of fluorescence in-situ hybridization mapping on rice mitotic prometaphase chromosomes, meiotic pachytene chromosomes and extended DNA fibers. Chromosome Res 10, 379-387. DOI URL
[8]	Do Vale Martins L, de Oliveira Bustamante F, da Silva Oliveira AR, da Costa AF, de Lima Feitoza L, Liang QH, Zhao HN, Benko-Iseppon AM, Muñoz-Amatriaín M, Pedrosa-Harand A, Jiang JM, Brasileiro-Vidal AC(2021). BAC- and Oligo-FISH mapping reveals chromosome evolution among Vigna angularis, V. unguiculata, and Phaseolus vulgaris. Chromosoma 130, 133-147. DOI PMID
[9]	Do Vale Martins L, Yu F, Zhao HN, Dennison T, Lauter N, Wang HY, Deng ZH, Thompson A, Semrau K, Rouillard JM, Birchler JA, Jiang JM (2019). Meiotic crossovers characterized by haplotype-specific chromosome painting in maize. Nat Commun 10, 4604. DOI PMID
[10]	Han YH, Zhang T, Thammapichai P, Weng YQ, Jiang JM (2015). Chromosome-specific painting in Cucumis species using bulked oligonucleotides. Genetics 200, 771-779. DOI URL
[11]	He L, Braz GT, Torres GA, Jiang JM (2018). Chromosome painting in meiosis reveals pairing of specific chromosomes in polyploid Solanum species. Chromosoma 127, 505-513. DOI
[12]	Hou LL, Xu M, Zhang T, Xu ZH, Wang WY, Zhang JX, Yu MM, Ji W, Zhu CW, Gong ZY, Gu MH, Jiang JM, Yu HX (2018). Chromosome painting and its applications in cultivated and wild rice. BMC Plant Biol 18, 110. DOI PMID
[13]	Idziak D, Betekhtin A, Wolny E, Lesniewska K, Wright J, Febrer M, Bevan MW, Jenkins G, Hasterok R (2011). Painting the chromosomes of Brachypodium: current status and future prospects. Chromosoma 120, 469-479. DOI URL
[14]	Jiang JM (2019). Fluorescence in situ hybridization in plants: recent developments and future applications. Chromosome Res 27, 153-165. DOI
[15]	Jiang JM, Gill BS (2006). Current status and the future of fluorescence in situ hybridization (FISH) in plant genome research. Genome 49, 1057-1068. DOI URL
[16]	Liu GQ, Zhang T (2021). Single copy oligonucleotide fluorescence in situ hybridization probe design platforms: development, application and evaluation. Int J Mol Sci 22, 7124. DOI URL
[17]	Liu XY, Sun S, Wu Y, Zhou Y, Gu SW, Yu HX, Yi CD, Gu MH, Jiang JM, Liu B, Zhang T, Gong ZY (2020). Dual-color Oligo-FISH can reveal chromosomal variations and evolution in Oryza species. Plant J 101, 112-121. DOI URL
[18]	Lou QF, Zhang YX, He YH, Li J, Jia L, Cheng CY, Guan W, Yang SQ, Chen JF (2014). Single-copy gene-based chromosome painting in cucumber and its application for chromosome rearrangement analysis in Cucumis. Plant J 78, 169-179. DOI URL
[19]	Lysak MA, Fransz PF, Ali HBM, Schubert I (2001). Chromosome painting in Arabidopsis thaliana. Plant J 28, 689-697. DOI PMID
[20]	Meng Z, Zhang ZL, Yan TY, Lin QF, Wang Y, Huang WY, Huang YJ, Li ZJ, Yu QY, Wang JP, Wang K (2018). Comprehensively characterizing the cytological features of Saccharum spontaneum by the development of a comp- lete set of chromosome-specific Oligo probes. Front Plant Sci 9, 1624. DOI URL
[21]	Qu MM, Li KP, Han YL, Chen L, Li ZY, Han YH (2017). Integrated karyotyping of woodland strawberry (Fragaria vesca) with Oligopaint FISH probes. Cytogenet Genome Res 153, 158-164. DOI PMID
[22]	Shi PY, Sun HJ, Liu GQ, Zhang X, Zhou JW, Song RR, Xiao J, Yuan CX, Sun L, Wang ZK, Lou QF, Jiang JM, Wang XE, Wang HY (2022). Chromosome painting reveals inter-chromosomal rearrangements and evolution of subgenome D of wheat. Plant J 112, 55-67. DOI URL
[23]	Xin HY, Zhang T, Han YH, Wu YF, Shi JS, Xi ML, Jiang JM (2018). Chromosome painting and comparative physical mapping of the sex chromosomes in Populus tomentosa and Populus deltoides. Chromosoma 127, 313-321. DOI
[24]	Yamada NA, Rector LS, Tsang P, Carr E, Scheffer A, Sederberg MC, Aston ME, Ach RA, Tsalenko A, Sampas N, Peter B, Bruhn L, Brothman AR (2011). Visualization of fine-scale genomic structure by oligonucleotide-based high-resolution FISH. Cytogenet Genome Res 132, 248-254. DOI PMID
[25]	Yu F, Zhao XW, Chai J, Ding XE, Li XT, Huang YJ, Wang XH, Wu JY, Zhang MQ, Yang QH, Deng ZH, Jiang JM (2022). Chromosome-specific painting unveils chromosomal fusions and distinct allopolyploid species in the Saccharum complex. New Phytol 233, 1953-1965. DOI URL
[26]	Zhang FF, Shen Y, Miao CB, Cao YW, Shi WQ, Du GJ, Tang D, Li YF, Luo Q, Cheng ZK (2020). OsRAD51D promotes homologous pairing and recombination by preventing nonhomologous interactions in rice meiosis. New Phytol 227, 824-839. DOI PMID
[27]	Zhang FF, Tang D, Shen Y, Xue ZH, Shi WQ, Ren LJ, Du GJ, Li YF, Cheng ZK (2017). The F-box protein ZYGO1 mediates bouquet formation to promote homologous pairing, synapsis, and recombination in rice meiosis. Plant Cell 29, 2597-2609. DOI URL
[28]	Zhang T, Liu GQ, Zhao HN, Braz GT, Jiang JM (2021). Chorus2: design of genome-scale oligonucleotide-based probes for fluorescence in situ hybridization. Plant Biotechnol J 19, 1967-1978. DOI PMID

植物寡核苷酸荧光原位杂交技术方法

Protocals for Oligonucleotide Fluorescence in situ Hybridization in Plants

RichHTML

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

图/表 2

参考文献 28

相关文章 15

编辑推荐

Metrics

本文评价

[1]	连佳丽, 陈婧, 杨雪琴, 赵莹, 罗叙, 韩翠, 赵雅欣, 李建平. 荒漠草原植物多样性和微生物多样性对降水变化的响应[J]. 生物多样性, 2024, 32(6): 24044-.
[2]	胡宗刚. 抗战胜利后中美曾筹划合编《中国植物志》[J]. 生物多样性, 2024, 32(6): 24220-.
[3]	何花, 谭敦炎, 杨晓琛. 被子植物隐型雌雄异株性系统的多样性、系统演化及进化意义[J]. 生物多样性, 2024, 32(6): 24149-.
[4]	江康威张青青王亚菲李宏丁雨杨永强吐尔逊娜依·热依木. 放牧干扰下天山北坡中段植物功能群特征及其与土壤环境因子的关系[J]. 植物生态学报, 2024, 48(6): 0-0.
[5]	蔡慧颖李兰慧林阳梁亚涛杨光孙龙. 白桦叶片和细根非结构性碳水化合物对火后时间的响应[J]. 植物生态学报, 2024, 48(6): 0-0.
[6]	巴苏艳, 赵春艳, 刘媛, 方强. 通过虫体花粉识别构建植物‒传粉者网络: 人工模型与AI模型高度一致[J]. 生物多样性, 2024, 32(6): 24088-.
[7]	艾妍雨, 胡海霞, 沈婷, 莫雨轩, 杞金华, 宋亮. 附生维管植物多样性及其与宿主特征的相关性: 以哀牢山中山湿性常绿阔叶林为例[J]. 生物多样性, 2024, 32(5): 24072-.
[8]	邓蓓王晓锋廖君. 环境胁迫影响三峡库区消落带草本和木本植物生理生态特征的整合分析[J]. 植物生态学报, 2024, 48(5): 623-637.
[9]	白皓然侯盟刘艳杰. 少花蒺藜草入侵与干旱对羊草草原生产力的影响机制[J]. 植物生态学报, 2024, 48(5): 577-589.
[10]	胡蝶蒋欣琪戴志聪陈戴一张雨祁珊珊杜道林. 丛枝菌根真菌提高入侵杂草南美蟛蜞菊对除草剂的耐受性[J]. 植物生态学报, 2024, 48(5): 651-659.
[11]	董劭琼, 侯东杰, 曲孝云, 郭柯. 柴达木盆地植物群落样方数据集[J]. 植物生态学报, 2024, 48(4): 534-540.
[12]	周玉滢, 陈辉, 刘斯穆. 植物非典型Aux/IAA蛋白应答生长素研究进展[J]. 植物学报, 2024, 59(4): 0-0.
[13]	曲泽坤, 朱丽琴, 姜琦, 王小红, 姚晓东, 蔡世锋, 罗素珍, 陈光水. 亚热带常绿阔叶林丛枝菌根树种养分觅食策略及其与细根形态间的关系[J]. 植物生态学报, 2024, 48(4): 416-427.
[14]	杨佳丽, 饶羽菲, 张润花, 周国林, 林处发, 何燕红, 宁国贵. 捕虫堇叶片高效再生体系的建立[J]. 植物学报, 2024, 59(4): 0-0.
[15]	付粱晨, 丁宗巨, 唐茂, 曾辉, 朱彪. 北京东灵山白桦和蒙古栎的根际效应及其季节动态[J]. 植物生态学报, 2024, 48(4): 508-522.