Cold Spring Harbor protocols最新文献

Detecting Meiotic Crossing-Overs in Maize Using Chromatin Immunoprecipitation-Sequencing (ChIP-seq). 利用染色质免疫沉淀测序（ChIP-seq）检测玉米减数分裂杂交。

Cold Spring Harbor protocols Pub Date : 2026-10-01 DOI: 10.1101/pdb.prot108741

Mateusz Zelkowski, Yan He, Adele Zhou, Wojciech P Pawlowski

{"title":"Detecting Meiotic Crossing-Overs in Maize Using Chromatin Immunoprecipitation-Sequencing (ChIP-seq).","authors":"Mateusz Zelkowski, Yan He, Adele Zhou, Wojciech P Pawlowski","doi":"10.1101/pdb.prot108741","DOIUrl":"10.1101/pdb.prot108741","url":null,"abstract":"During meiosis, homologous chromosomes engage in reciprocal exchanges of segments in a process known as crossing over (CO). About 85% of CO events in maize are products of the class I pathway. Class I COs are interference-sensitive, meaning that the formation of one CO reduces the likelihood of another CO forming close by. This protocol describes a chromatin immunoprecipitation-sequencing (ChIP-seq)-based method for mapping meiotic COs in maize, using an antibody against MutL Homolog 3 (MLH3), a key component of the class I CO pathway. CO sites are determined by Illumina sequencing of DNA isolated from MLH3-associated chromatin fragments. Traditionally, COs have been identified through genetic mapping, which relies on the segregation of genetic markers in the progeny of hybrid plants. However, conventional genetic mapping provides limited resolution and requires large numbers of progeny individuals. The MLH3 ChIP-seq approach enables direct detection of COs, providing high-resolution and genome-wide coverage, including genome regions with low DNA sequence polymorphism, which are inaccessible to genetic CO mapping. Furthermore, MLH3 ChIP-seq enables screening of thousands of CO events, greatly accelerating the analysis and reducing its cost. This protocol can also be used to examine any chromatin-bound meiotic proteins and adapted to studying chromatin-associated proteins in somatic cells.","PeriodicalId":10496,"journal":{"name":"Cold Spring Harbor protocols","volume":" ","pages":"pdb.prot108741"},"PeriodicalIF":0.0,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148928915","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Analyzing Meiosis in Maize. 玉米减数分裂分析。

Cold Spring Harbor protocols Pub Date : 2026-10-01 DOI: 10.1101/pdb.top108472

Olga Zimina, Juli Jing, Mateusz Zelkowski, Wojciech P Pawlowski

{"title":"Analyzing Meiosis in Maize.","authors":"Olga Zimina, Juli Jing, Mateusz Zelkowski, Wojciech P Pawlowski","doi":"10.1101/pdb.top108472","DOIUrl":"10.1101/pdb.top108472","url":null,"abstract":"Meiosis is central to sexual reproduction and the main source of genetic diversity in plants. Understanding how meiotic processes are regulated has direct relevance to agriculture. As meiotic recombination is the vehicle of plant breeding, gaining the ability to influence recombination patterns can accelerate crop improvement. Maize is a powerful model for studying plant meiosis, thanks to its large chromosomes, well-developed genetics, and the availability of diverse cytogenetic and molecular tools. Insights gained from maize studies can extend to other species. In this review, we describe a variety of approaches for examining meiosis and meiotic recombination in maize. Cytological techniques, including protein immunolocalization and fluorescence in situ hybridization (FISH), enable visualization of chromosome structure and behavior, as well as crossover (CO) formation. Chromatin immunoprecipitation (ChIP) is used in meiosis research to determine locations of recombination proteins, identify recombination sites, and elucidate chromatin features, such as histone modifications. Quantification of COs at specific genomic sites through pollen typing by droplet digital PCR allows precise high-resolution measurement of recombination rates. Combining cytology, protein localization, and molecular assays provides a multiscale picture of meiosis, linking molecular mechanisms to chromosome behavior and, ultimately, to genetic variation.","PeriodicalId":10496,"journal":{"name":"Cold Spring Harbor protocols","volume":" ","pages":"pdb.top108472"},"PeriodicalIF":0.0,"publicationDate":"2026-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148928772","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Cytological Assessment of Maize Pollen Viability Using a Simplified Staining Protocol. 用简化染色法评价玉米花粉活力的细胞学研究。

Cold Spring Harbor protocols Pub Date : 2026-10-01 DOI: 10.1101/pdb.prot108742

Mateusz Zelkowski, Wojciech P Pawlowski

引用次数: 0

Corrigendum: Generation of a Phage Display Chicken Single-Chain Variable Fragment Library. 更正：噬菌体展示鸡单链可变片段文库的生成。

Cold Spring Harbor protocols Pub Date : 2026-10-01 DOI: 10.1101/pdb.corr108750

Hyunji Yang, Jisu Chae, Hyori Kim, Jinsung Noh, Junho Chung

引用次数: 0

Site-Specific Measurement of Meiotic Crossing-Over Rate with Droplet Digital PCR. 用微滴数字PCR测定减数分裂交叉率的位点特异性研究。

Cold Spring Harbor protocols Pub Date : 2026-10-01 DOI: 10.1101/pdb.prot108743

Olga Zimina, Mateusz Zelkowski, Andrew Ziesel, Wojciech P Pawlowski

引用次数: 0

Immunolocalization of Meiotic Proteins in Maize. 玉米减数分裂蛋白的免疫定位。

Cold Spring Harbor protocols Pub Date : 2026-10-01 DOI: 10.1101/pdb.prot108739

Juli Jing, Wojciech P Pawlowski

引用次数: 0

Preparation of Meiotic Chromosome Spreads and Fluorescence In Situ Hybridization of Maize Meiotic Chromosomes. 玉米减数分裂染色体扩增的制备及染色体荧光原位杂交。

Cold Spring Harbor protocols Pub Date : 2026-10-01 DOI: 10.1101/pdb.prot108738

Juli Jing, Wojciech P Pawlowski

引用次数: 0

Immuno-FISH: Simultaneous Chromosome Painting and Immunostaining of Proteins in Maize Meiocytes. 免疫- fish：玉米减数细胞中蛋白质的同步染色体染色和免疫染色。

Cold Spring Harbor protocols Pub Date : 2026-10-01 DOI: 10.1101/pdb.prot108740

Olga Zimina, Wojciech P Pawlowski

引用次数: 0

Methyl-Salicylate Clearing and Imaging of Maize Meristems. 玉米分生组织中水杨酸甲酯的清除与成像。

Cold Spring Harbor protocols Pub Date : 2026-09-21 DOI: 10.1101/pdb.prot108749

Samuel Leiboff, Annis E Richardson

{"title":"Methyl-Salicylate Clearing and Imaging of Maize Meristems.","authors":"Samuel Leiboff, Annis E Richardson","doi":"10.1101/pdb.prot108749","DOIUrl":"10.1101/pdb.prot108749","url":null,"abstract":"The final morphology of a plant is determined by much earlier events that occur during development. Meristems are critical for the development of the aerial parts of plants, such as leaves and stems. The morphology of these meristems, which can vary significantly across genetic backgrounds, correlates with phenotypic traits in Zea mays (maize) that influence yield, such as leaf initiation rate and kernel row number. Traditional methods for morphological analysis of meristems, including histological sectioning and fine dissection, often require considerable technical expertise. To address these challenges, we present a detailed protocol employing methyl-salicylate for tissue clearing, combined with Differential Interference Contrast (DIC) microscopy to visualize maize shoot apical meristems. This method enables the intact visualization of vegetative meristems without the need for extensive dissection, thereby preserving the natural morphology of the meristem. Using this approach, the meristem is coarsely dissected, fixed, optionally stained, and cleared to enable visualization through leaf layers. By improving the accessibility and quality of meristem imaging, this protocol facilitates accurate quantification of meristem morphology, critical for genetic and developmental studies. Optimized primarily for maize, the outlined techniques can also be adapted for other plant species, providing a versatile tool in the field of plant developmental biology.","PeriodicalId":10496,"journal":{"name":"Cold Spring Harbor protocols","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2026-09-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148929049","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Fresh Tissue Scanning Electron Microscopy (SEM) of Maize Meristems. 玉米分生组织的新鲜组织扫描电镜（SEM）。

Cold Spring Harbor protocols Pub Date : 2026-09-10 DOI: 10.1101/pdb.prot108748

Comfort Bonney Arku, Madelaine Bartlett, Annis E Richardson

{"title":"Fresh Tissue Scanning Electron Microscopy (SEM) of Maize Meristems.","authors":"Comfort Bonney Arku, Madelaine Bartlett, Annis E Richardson","doi":"10.1101/pdb.prot108748","DOIUrl":"https://doi.org/10.1101/pdb.prot108748","url":null,"abstract":"In plants, final organ shape and position influence function and whole plant productivity. In Zea mays (maize), leaf, ear, and tassel morphology are all key agronomic features influencing yield. These aerial structures originate from stem cell populations within meristems. Understanding how meristems are regulated and how stem cells differentiate over developmental time to form mature leaves, ears, and tassels is important for identifying novel routes for maize improvement. High-resolution imaging of meristem morphology is a key component of developmental studies. Scanning electron microscopy (SEM) is widely used to visualize meristem topology; however, conventional SEM preparation requires fixation, dehydration, drying, and conductive coating. These steps are time-consuming, technically demanding, and can introduce damage. Here, we present a rapid protocol for imaging freshly dissected meristems from maize using a benchtop scanning electron microscope. Fresh tissues are mounted directly onto conductive adhesive and imaged under high vacuum at low accelerating voltage, enabling high-quality micrographs to be captured within minutes of dissection. Although dehydration and tissue collapse can occur under vacuum, rapid sample handling and image acquisition minimize these effects and produce images comparable to those obtained from conventionally prepared specimens. This streamlined workflow substantially reduces preparation time, lowers material requirements, and facilitates medium-throughput morphological analysis. The method provides a practical and reproducible approach for rapid developmental phenotyping and is readily adaptable to other plant species and other tissues.","PeriodicalId":10496,"journal":{"name":"Cold Spring Harbor protocols","volume":" ","pages":""},"PeriodicalIF":0.0,"publicationDate":"2026-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148929068","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0