Applications in Plant Sciences最新文献

{"title":"Correction to “A comparison of freezer-stored DNA and herbarium tissue samples for chloroplast assembly and genome skimming”","authors":"","doi":"10.1002/aps3.11540","DOIUrl":"10.1002/aps3.11540","url":null,"abstract":"<p>McAssey, E. V., Downs, C., Yorkston, M., Morden, C., and Heyduk, K. 2023. A comparison of freezer-stored DNA and herbarium tissue samples for chloroplast assembly and genome skimming. <i>Applications in Plant Sciences</i> 11(3): e11527</p><p>A statistical error was found after article publication. The relevant text from the Results section is provided below, with the corrected values shown in bold text. The error does not affect the findings of the study.</p><p>“Herbarium tissue library samples had significantly smaller insert sizes of mapped chloroplast reads compared to their freezer-stored DNA paired samples, taking into account covariates of read numbers and year (<b><i>F</i>_1,25 = 229.243</b>, <i><b>P</b></i> &lt; <b>0.001</b>). There was also a significant interaction effect between library size and sampling year (<b><i>F</i>_1,25 = 9.753</b>, <i>P</i> &lt; 0.01). Similarly, herbarium tissue samples also had higher amounts of adapter sequences in the reads (<b><i>F</i>_1,25 = 85.009</b>, <i>P</i> &lt; 0.001), with sampling year a significant covariate in the model (<b><i>F</i>_1,25 = 6.378</b>, <i>P</i> &lt; 0.05).”</p><p>We apologize for this error.</p>","PeriodicalId":8022,"journal":{"name":"Applications in Plant Sciences","volume":"11 4","pages":""},"PeriodicalIF":3.6,"publicationDate":"2023-08-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10439819/pdf/APS3-11-e11540.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10034571","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"GOgetter: A pipeline for summarizing and visualizing GO slim annotations for plant genetic data","authors":"Emily B. Sessa,&nbsp;Rishi R. Masalia,&nbsp;Nils Arrigo,&nbsp;Michael S. Barker,&nbsp;Jessie A. Pelosi","doi":"10.1002/aps3.11536","DOIUrl":"10.1002/aps3.11536","url":null,"abstract":"<div>\u0000 \u0000 \u0000 <section>\u0000 \u0000 <h3> Premise</h3>\u0000 \u0000 <p>The functional annotation of genes is a crucial component of genomic analyses. A common way to summarize functional annotations is with hierarchical gene ontologies, such as the Gene Ontology (GO) Resource. GO includes information about the cellular location, molecular function(s), and products/processes that genes produce or are involved in. For a set of genes, summarizing GO annotations using pre-defined, higher-order terms (GO slims) is often desirable in order to characterize the overall function of the data set, and it is impractical to do this manually.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Methods and Results</h3>\u0000 \u0000 <p>The GOgetter pipeline consists of bash and Python scripts. From an input FASTA file of nucleotide gene sequences, it outputs text and image files that list (1) the best hit for each input gene in a set of reference gene models, (2) all GO terms and annotations associated with those hits, and (3) a summary and visualization of GO slim categories for the data set. These output files can be queried further and analyzed statistically, depending on the downstream need(s).</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Conclusions</h3>\u0000 \u0000 <p>GO annotations are a widely used “universal language” for describing gene functions and products. GOgetter is a fast and easy-to-implement pipeline for obtaining, summarizing, and visualizing GO slim categories associated with a set of genes.</p>\u0000 </section>\u0000 </div>","PeriodicalId":8022,"journal":{"name":"Applications in Plant Sciences","volume":"11 4","pages":""},"PeriodicalIF":3.6,"publicationDate":"2023-08-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://bsapubs.onlinelibrary.wiley.com/doi/epdf/10.1002/aps3.11536","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10052130","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Target capture and genome skimming for plant diversity studies","authors":"Flávia Fonseca Pezzini,&nbsp;Giada Ferrari,&nbsp;Laura L. Forrest,&nbsp;Michelle L. Hart,&nbsp;Kanae Nishii,&nbsp;Catherine A. Kidner","doi":"10.1002/aps3.11537","DOIUrl":"10.1002/aps3.11537","url":null,"abstract":"<p>Recent technological advances in long-read high-throughput sequencing and assembly methods have facilitated the generation of annotated chromosome-scale whole-genome sequence data for evolutionary studies; however, generating such data can still be difficult for many plant species. For example, obtaining high-molecular-weight DNA is typically impossible for samples in historical herbarium collections, which often have degraded DNA. The need to fast-freeze newly collected living samples to conserve high-quality DNA can be complicated when plants are only found in remote areas. Therefore, short-read reduced-genome representations, such as target capture and genome skimming, remain important for evolutionary studies. Here, we review the pros and cons of each technique for non-model plant taxa. We provide guidance related to logistics, budget, the genomic resources previously available for the target clade, and the nature of the study. Furthermore, we assess the available bioinformatic analyses, detailing best practices and pitfalls, and suggest pathways to combine newly generated data with legacy data. Finally, we explore the possible downstream analyses allowed by the type of data generated using each technique. We provide a practical guide to help researchers make the best-informed choice regarding reduced genome representation for evolutionary studies of non-model plants in cases where whole-genome sequencing remains impractical.</p>","PeriodicalId":8022,"journal":{"name":"Applications in Plant Sciences","volume":"11 4","pages":""},"PeriodicalIF":3.6,"publicationDate":"2023-08-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/bb/3e/APS3-11-e11537.PMC10439825.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10356348","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Welcome to the big leaves: Best practices for improving genome annotation in non-model plant genomes","authors":"Vidya S. Vuruputoor,&nbsp;Daniel Monyak,&nbsp;Karl C. Fetter,&nbsp;Cynthia Webster,&nbsp;Akriti Bhattarai,&nbsp;Bikash Shrestha,&nbsp;Sumaira Zaman,&nbsp;Jeremy Bennett,&nbsp;Susan L. McEvoy,&nbsp;Madison Caballero,&nbsp;Jill L. Wegrzyn","doi":"10.1002/aps3.11533","DOIUrl":"10.1002/aps3.11533","url":null,"abstract":"<div>\u0000 \u0000 \u0000 <section>\u0000 \u0000 <h3> Premise</h3>\u0000 \u0000 <p>Robust standards to evaluate quality and completeness are lacking in eukaryotic structural genome annotation, as genome annotation software is developed using model organisms and typically lacks benchmarking to comprehensively evaluate the quality and accuracy of the final predictions. The annotation of plant genomes is particularly challenging due to their large sizes, abundant transposable elements, and variable ploidies. This study investigates the impact of genome quality, complexity, sequence read input, and method on protein-coding gene predictions.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Methods</h3>\u0000 \u0000 <p>The impact of repeat masking, long-read and short-read inputs, and de novo and genome-guided protein evidence was examined in the context of the popular BRAKER and MAKER workflows for five plant genomes. The annotations were benchmarked for structural traits and sequence similarity.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Results</h3>\u0000 \u0000 <p>Benchmarks that reflect gene structures, reciprocal similarity search alignments, and mono-exonic/multi-exonic gene counts provide a more complete view of annotation accuracy. Transcripts derived from RNA-read alignments alone are not sufficient for genome annotation. Gene prediction workflows that combine evidence-based and ab initio approaches are recommended, and a combination of short and long reads can improve genome annotation. Adding protein evidence from de novo assemblies, genome-guided transcriptome assemblies, or full-length proteins from OrthoDB generates more putative false positives as implemented in the current workflows. Post-processing with functional and structural filters is highly recommended.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Discussion</h3>\u0000 \u0000 <p>While the annotation of non-model plant genomes remains complex, this study provides recommendations for inputs and methodological approaches. We discuss a set of best practices to generate an optimal plant genome annotation and present a more robust set of metrics to evaluate the resulting predictions.</p>\u0000 </section>\u0000 </div>","PeriodicalId":8022,"journal":{"name":"Applications in Plant Sciences","volume":"11 4","pages":""},"PeriodicalIF":3.6,"publicationDate":"2023-08-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://bsapubs.onlinelibrary.wiley.com/doi/epdf/10.1002/aps3.11533","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10050062","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"hybpiper-nf and paragone-nf: Containerization and additional options for target capture assembly and paralog resolution","authors":"Chris Jackson,&nbsp;Todd McLay,&nbsp;Alexander N. Schmidt-Lebuhn","doi":"10.1002/aps3.11532","DOIUrl":"10.1002/aps3.11532","url":null,"abstract":"<div>\u0000 \u0000 \u0000 <section>\u0000 \u0000 <h3> Premise</h3>\u0000 \u0000 <p>The HybPiper pipeline has become one of the most widely used tools for the assembly of target capture data for phylogenomic analysis. After the production of locus sequences and before phylogenetic analysis, the identification of paralogs is a critical step for ensuring the accurate inference of evolutionary relationships. Algorithmic approaches using gene tree topologies for the inference of ortholog groups are computationally efficient and broadly applicable to non-model organisms, especially in the absence of a known species tree.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Methods and Results</h3>\u0000 \u0000 <p>We containerized and expanded the functionality of both HybPiper and a pipeline for the inference of ortholog groups, providing novel options for the treatment of target capture sequence data, and allowing seamless use of the outputs of the former as inputs for the latter. The Singularity container presented here includes all dependencies, and the corresponding pipelines (hybpiper-nf and paragone-nf, respectively) are implemented via two Nextflow scripts for easier deployment and to vastly reduce the number of commands required for their use.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Conclusions</h3>\u0000 \u0000 <p>The hybpiper-nf and paragone-nf pipelines are easily installed and provide a user-friendly experience and robust results to the phylogenetic community. They are used by the Australian Angiosperm Tree of Life project. The pipelines are available at https://github.com/chrisjackson-pellicle/hybpiper-nf and https://github.com/chrisjackson-pellicle/paragone-nf.</p>\u0000 </section>\u0000 </div>","PeriodicalId":8022,"journal":{"name":"Applications in Plant Sciences","volume":"11 4","pages":""},"PeriodicalIF":3.6,"publicationDate":"2023-07-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/f4/1c/APS3-11-e11532.PMC10439820.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10046655","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A field-capable rapid plant DNA extraction protocol using microneedle patches for botanical surveying and monitoring","authors":"Jonathan Selz,&nbsp;Nicolas R. Adam,&nbsp;Céline E. M. Magrini,&nbsp;Fulvia Malvido Montandon,&nbsp;Sven Buerki,&nbsp;Sebastian J. Maerkl","doi":"10.1002/aps3.11529","DOIUrl":"10.1002/aps3.11529","url":null,"abstract":"<div>\u0000 \u0000 \u0000 <section>\u0000 \u0000 <h3> Premise</h3>\u0000 \u0000 <p>A novel protocol for rapid plant DNA extraction using microneedles is proposed, which supports botanic surveys, taxonomy, and systematics. This protocol can be conducted in the field with limited laboratory skills and equipment. The protocol is validated by sequencing and comparing the results with QIAGEN spin-column DNA extractions using BLAST analyses.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Methods and Results</h3>\u0000 \u0000 <p>Two sets of DNA extractions were conducted on 13 species spanning various leaf anatomies and phylogenetic lineages: (i) fresh leaves were punched with custom polymeric microneedle patches to recover genomic DNA, or (ii) QIAGEN DNA extractions. Three plastid (<i>matK</i>, <i>rbcL</i>, and <i>trnH-psbA</i>) and one nuclear ribosomal (ITS) DNA regions were amplified and sequenced using Sanger or nanopore technology. The proposed method reduced the extraction time to 1 min and yielded the same DNA sequences as the QIAGEN extractions.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Conclusions</h3>\u0000 \u0000 <p>Our drastically faster and simpler method is compatible with nanopore sequencing and is suitable for multiple applications, including high-throughput DNA-based species identifications and monitoring.</p>\u0000 </section>\u0000 </div>","PeriodicalId":8022,"journal":{"name":"Applications in Plant Sciences","volume":"11 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2023-06-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://bsapubs.onlinelibrary.wiley.com/doi/epdf/10.1002/aps3.11529","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9764040","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Modified CTAB protocols for high-molecular-weight DNA extractions from ferns","authors":"Pei-Jun Xie,&nbsp;Ya-Ting Ke,&nbsp;Li-Yaung Kuo","doi":"10.1002/aps3.11526","DOIUrl":"10.1002/aps3.11526","url":null,"abstract":"<div>\u0000 \u0000 \u0000 <section>\u0000 \u0000 <h3> Premise</h3>\u0000 \u0000 <p>Efficient protocols for extracting high-molecular-weight (HMW) DNA from ferns facilitate the long-read sequencing of their large and complex genomes. Here, we perform two cetyltrimethylammonium bromide (CTAB)-based protocols to extract HMW DNA and evaluate their applicability in diverse fern taxa for the first time.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Methods and Results</h3>\u0000 \u0000 <p>We describe two modified CTAB protocols, with key adjustments to minimize mechanical disruption during lysis to prevent DNA shearing. One of these protocols uses a small amount of fresh tissue but yields a considerable quantity of HMW DNA with high efficiency. The other accommodates a large amount of input tissue, adopts an initial step of nuclei isolation, and thus ensures a high yield in a short period of time. Both methods were proven to be robust and effective in obtaining HMW DNA from diverse fern lineages, including 33 species in 19 families. The DNA extractions mostly had high DNA integrity, with mean sizes larger than 50 kbp, as well as high purity (A₂₆₀/A₂₃₀ and A₂₆₀/A₂₈₀ &gt; 1.8).</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Conclusions</h3>\u0000 \u0000 <p>This study provides HMW DNA extraction protocols for ferns in the hope of facilitating further attempts to sequence their genomes, which will bridge our genomic understanding of land plant diversity.</p>\u0000 </section>\u0000 </div>","PeriodicalId":8022,"journal":{"name":"Applications in Plant Sciences","volume":"11 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2023-06-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/32/c2/APS3-11-e11526.PMC10278929.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9709978","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"High-molecular-weight DNA extraction for long-read sequencing of plant genomes: An optimization of standard methods","authors":"Myoungbo Kang,&nbsp;Andre Chanderbali,&nbsp;Seungyeon Lee,&nbsp;Douglas E. Soltis,&nbsp;Pamela S. Soltis,&nbsp;Sangtae Kim","doi":"10.1002/aps3.11528","DOIUrl":"10.1002/aps3.11528","url":null,"abstract":"<div>\u0000 \u0000 \u0000 <section>\u0000 \u0000 <h3> Premise</h3>\u0000 \u0000 <p>Developing an effective and easy-to-use high-molecular-weight (HMW) DNA extraction method is essential for genomic research, especially in the era of third-generation sequencing. To efficiently use technologies capable of generating long-read sequences, it is important to maximize both the length and purity of the extracted DNA; however, this is frequently difficult to achieve with plant samples.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Methods and Results</h3>\u0000 \u0000 <p>We present a HMW DNA extraction method that combines (1) a nuclei extraction method followed by (2) a traditional cetyltrimethylammonium bromide (CTAB) DNA extraction method for plants with optimized extraction conditions that influence HMW DNA recovery. Our protocol produced DNA fragments (percentage of fragments &gt;20 kbp) that were, on average, ca. five times longer than those obtained using a commercial kit, and contaminants were removed more effectively.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Conclusions</h3>\u0000 \u0000 <p>This effective HMW DNA extraction protocol can be used as a standard protocol for a diverse array of taxa, which will enhance plant genomic research.</p>\u0000 </section>\u0000 </div>","PeriodicalId":8022,"journal":{"name":"Applications in Plant Sciences","volume":"11 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2023-06-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://bsapubs.onlinelibrary.wiley.com/doi/epdf/10.1002/aps3.11528","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9764041","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A metabarcoding protocol targeting two DNA regions to analyze root-associated fungal communities in ferns and lycophytes","authors":"Thais Guillen-Otero,&nbsp;Soon-Jae Lee,&nbsp;Cheng-Wei Chen,&nbsp;Peter Szoevenyi,&nbsp;Michael Kessler","doi":"10.1002/aps3.11523","DOIUrl":"10.1002/aps3.11523","url":null,"abstract":"<div>\u0000 \u0000 \u0000 <section>\u0000 \u0000 <h3> Premise</h3>\u0000 \u0000 <p>Detailed studies of the fungi associated with lycophytes and ferns provide crucial insights into the early evolution of land plants. However, most investigations to date have assessed fern–fungus interactions based only on visual root inspection. In the present research, we establish and evaluate a metabarcoding protocol to analyze the fungal communities associated with fern and lycophyte roots.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Methods</h3>\u0000 \u0000 <p>We used two primer pairs focused on the ITS rRNA region to screen the general fungal communities, and the 18S rRNA to target Glomeromycota fungi (i.e., arbuscular mycorrhizal fungi). To test these approaches, we collected and processed roots from 12 phylogenetically distant fern and lycophyte species.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Results</h3>\u0000 \u0000 <p>We found marked compositional differences between the ITS and 18S data sets. While the ITS data set demonstrated the dominance of orders Glomerales (phylum Glomeromycota), Pleosporales, and Helotiales (both in phylum Ascomycota), the 18S data set revealed the greatest diversity of Glomeromycota. Non-metric multidimensional scaling (NMDS) ordination suggested an important geographical effect in sample similarities.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Discussion</h3>\u0000 \u0000 <p>The ITS-based approach is a reliable and effective method to analyze the fungal communities associated with fern and lycophyte roots. The 18S approach is more appropriate for studies focused on the detailed screening of arbuscular mycorrhizal fungi.</p>\u0000 </section>\u0000 </div>","PeriodicalId":8022,"journal":{"name":"Applications in Plant Sciences","volume":"11 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2023-06-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/53/3c/APS3-11-e11523.PMC10278937.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9712585","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Balancing read length and sequencing depth: Optimizing Nanopore long-read sequencing for monocots with an emphasis on the Liliales","authors":"Gisel Y. De La Cerda,&nbsp;Jacob B. Landis,&nbsp;Evan Eifler,&nbsp;Adriana I. Hernandez,&nbsp;Fay-Wei Li,&nbsp;Jing Zhang,&nbsp;Carrie M. Tribble,&nbsp;Nisa Karimi,&nbsp;Patricia Chan,&nbsp;Thomas Givnish,&nbsp;Susan R. Strickler,&nbsp;Chelsea D. Specht","doi":"10.1002/aps3.11524","DOIUrl":"10.1002/aps3.11524","url":null,"abstract":"<div>\u0000 \u0000 \u0000 <section>\u0000 \u0000 <h3> Premise</h3>\u0000 \u0000 <p>We present approaches used to generate long-read Nanopore sequencing reads for the Liliales and demonstrate how modifications to standard protocols directly impact read length and total output. The goal is to help those interested in generating long-read sequencing data determine which steps may be necessary for optimizing output and results.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Methods</h3>\u0000 \u0000 <p>Four species of <i>Calochortus</i> (Liliaceae) were sequenced. Modifications made to sodium dodecyl sulfate (SDS) extractions and cleanup protocols included grinding with a mortar and pestle, using cut or wide-bore tips, chloroform cleaning, bead cleaning, eliminating short fragments, and using highly purified DNA.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Results</h3>\u0000 \u0000 <p>Steps taken to maximize read length can decrease overall output. Notably, the number of pores in a flow cell is correlated with the overall output, yet we did not see an association between the pore number and the read length or the number of reads produced.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Discussion</h3>\u0000 \u0000 <p>Many factors contribute to the overall success of a Nanopore sequencing run. We showed the direct impact that several modifications to the DNA extraction and cleaning steps have on the total sequencing output, read size, and number of reads generated. We show a tradeoff between read length and the number of reads and, to a lesser extent, the total sequencing output, all of which are important factors for successful de novo genome assembly.</p>\u0000 </section>\u0000 </div>","PeriodicalId":8022,"journal":{"name":"Applications in Plant Sciences","volume":"11 3","pages":""},"PeriodicalIF":3.6,"publicationDate":"2023-06-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://bsapubs.onlinelibrary.wiley.com/doi/epdf/10.1002/aps3.11524","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9712582","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}