scholarly journals A Simple Genome Walking Strategy to Isolate Unknown Genomic Regions Using Long Primer and RAPD Primer

2019 ◽  
Vol 17 (2) ◽  
pp. 89-93
Author(s):  
Fajun Li ◽  
Chunpeng Fu ◽  
Qunfeng Li
Keyword(s):  
Genome Walking ◽  
Rapd Primer ◽  
Genomic Regions

Isolating promoters of multigene family members from the polyploid sugarcane genome by PCR-based walking in BAC DNA

Genome ◽  
2010 ◽  
Vol 53 (10) ◽  
pp. 840-847 ◽  
Author(s):  
Mona B. Damaj ◽  
Phillip D. Beremand ◽  
Marco T. Buenrostro-Nava ◽  
John Ivy ◽  
Siva P. Kumpatla ◽  
...  
Keyword(s):  
Multigene Family ◽  
Family Members ◽  
Genome Walking ◽  
Cdna Sequences ◽  
Regulated Expression ◽  
Specific Amplification ◽  
Polyploid Genome ◽  
Polymerase Chain ◽  
Genomic Regions ◽  
Target Promoter

The availability of a wider range of promoters for regulated expression in valuable transgenic crops would benefit functional genomics studies and current biotechnology programs aimed at improved productivity. Polymerase chain reaction (PCR)-based genome walking techniques are commonly used to isolate promoters or 5′ flanking genomic regions adjacent to known cDNA sequences in genomes that are not yet completely sequenced. However, these techniques are problematic when applied directly to DNA isolated from crops with highly complex and large genomes. An adaptor ligation-mediated PCR-based BAC genome walking method is described here for the efficient isolation of promoters of multigene family members, such as the putative defense and fiber biosynthesis DIRIGENT genes that are abundant in the stem of sugarcane, a species with a highly polyploid genome. The advantage of this method is the efficient and specific amplification of the target promoter using BAC genomic DNA as template for the adaptor ligation-mediated PCR walking.

Genetic Effects of Key Genomic Regions Controlling Yield-Related Traits in Wheat Founder Parent Fan 6

2018 ◽  
Vol 44 (5) ◽  
pp. 706
Author(s):  
Mei DENG ◽  
Yuan-Jiang HE ◽  
Lu-Lu GOU ◽  
Fang-Jie YAO ◽  
Jian LI ◽  
...  
Keyword(s):  
Genetic Effects ◽  
Genomic Regions ◽  
Founder Parent

Identification of genomic islands in Synechococcus sp. WH8102 using genomic barcode and whole-genome microarray analyses

2020 ◽  
Vol 15 ◽  
Author(s):  
Jiahui Pan ◽  
Xizi Luo ◽  
Tong Shao ◽  
Chaoying Li ◽  
Tingting Zhao ◽  
...  
Keyword(s):  
Genomic Islands ◽  
Whole Genome ◽  
Photosynthetic Organisms ◽  
Depth Study ◽  
Whole Genome Microarray ◽  
Integrated Methods ◽  
Synechococcus Sp ◽  
Intuitive View ◽  
Genomic Regions ◽  
Different Origins

Background: Synechococcus sp. WH8102 is one of the most abundant photosynthetic organisms in many ocean regions. Objective: The aim of this study is to identify genomic islands (GIs) in Synechococcus sp. WH8102 with integrated methods. Methods: We have applied genomic barcode to identify the GIs in Synechococcus sp. WH8102, which could make genomic regions of different origins visually apparent. The gene expression data of the predicted GIs was analyzed through microarray data which was collected for functional analysis of the relevant genes. Results: Seven GIs were identified in Synechococcus sp. WH8102. Most of them are involved in cell surface modification, photosynthesis and drug resistance. In addition, our analysis also revealed the functions of these GIs, which could be used for in-depth study on the evolution of this strain. Conclusion: Genomic barcodes provide us with a comprehensive and intuitive view of the target genome. We can use it to understand the intrinsic characteristics of the whole genome and identify GIs or other similar elements.

scholarly journals Assessing genomic diversity and signatures of selection in Jiaxian Red cattle using whole-genome sequencing data

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Xiaoting Xia ◽  
Shunjin Zhang ◽  
Huaju Zhang ◽  
Zijing Zhang ◽  
Ningbo Chen ◽  
...  
Keyword(s):  
Population Structure ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Genomic Variation ◽  
Genomic Diversity ◽  
System Response ◽  
Whole Genome ◽  
Population Structure Analysis ◽  
Native Cattle ◽  
Genomic Regions

Abstract Background Native cattle breeds are an important source of genetic variation because they might carry alleles that enable them to adapt to local environment and tough feeding conditions. Jiaxian Red, a Chinese native cattle breed, is reported to have originated from crossbreeding between taurine and indicine cattle; their history as a draft and meat animal dates back at least 30 years. Using whole-genome sequencing (WGS) data of 30 animals from the core breeding farm, we investigated the genetic diversity, population structure and genomic regions under selection of Jiaxian Red cattle. Furthermore, we used 131 published genomes of world-wide cattle to characterize the genomic variation of Jiaxian Red cattle. Results The population structure analysis revealed that Jiaxian Red cattle harboured the ancestry with East Asian taurine (0.493), Chinese indicine (0.379), European taurine (0.095) and Indian indicine (0.033). Three methods (nucleotide diversity, linkage disequilibrium decay and runs of homozygosity) implied the relatively high genomic diversity in Jiaxian Red cattle. We used θπ, CLR, FST and XP-EHH methods to look for the candidate signatures of positive selection in Jiaxian Red cattle. A total number of 171 (θπ and CLR) and 17 (FST and XP-EHH) shared genes were identified using different detection strategies. Functional annotation analysis revealed that these genes are potentially responsible for growth and feed efficiency (CCSER1), meat quality traits (ROCK2, PPP1R12A, CYB5R4, EYA3, PHACTR1), fertility (RFX4, SRD5A2) and immune system response (SLAMF1, CD84 and SLAMF6). Conclusion We provide a comprehensive overview of sequence variations in Jiaxian Red cattle genomes. Selection signatures were detected in genomic regions that are possibly related to economically important traits in Jiaxian Red cattle. We observed a high level of genomic diversity and low inbreeding in Jiaxian Red cattle. These results provide a basis for further resource protection and breeding improvement of this breed.

scholarly journals panRGP: a pangenome-based method to predict genomic islands and explore their diversity

2020 ◽  
Vol 36 (Supplement_2) ◽  
pp. i651-i658 ◽  
Author(s):  
Adelme Bazin ◽  
Guillaume Gautreau ◽  
Claudine Médigue ◽  
David Vallenet ◽  
Alexandra Calteau
Keyword(s):  
Escherichia Coli ◽  
Gene Transfer ◽  
Horizontal Gene Transfer ◽  
Species Level ◽  
Genomic Islands ◽  
Genome Plasticity ◽  
Reference Dataset ◽  
Prokaryotic Genomes ◽  
Ideal Approach ◽  
Genomic Regions

Abstract Motivation Horizontal gene transfer (HGT) is a major source of variability in prokaryotic genomes. Regions of genome plasticity (RGPs) are clusters of genes located in highly variable genomic regions. Most of them arise from HGT and correspond to genomic islands (GIs). The study of those regions at the species level has become increasingly difficult with the data deluge of genomes. To date, no methods are available to identify GIs using hundreds of genomes to explore their diversity. Results We present here the panRGP method that predicts RGPs using pangenome graphs made of all available genomes for a given species. It allows the study of thousands of genomes in order to access the diversity of RGPs and to predict spots of insertions. It gave the best predictions when benchmarked along other GI detection tools against a reference dataset. In addition, we illustrated its use on metagenome assembled genomes by redefining the borders of the leuX tRNA hotspot, a well-studied spot of insertion in Escherichia coli. panRPG is a scalable and reliable tool to predict GIs and spots making it an ideal approach for large comparative studies. Availability and implementation The methods presented in the current work are available through the following software: https://github.com/labgem/PPanGGOLiN. Detailed results and scripts to compute the benchmark metrics are available at https://github.com/axbazin/panrgp_supdata.

scholarly journals Inhibition of CRISPR-Cas12a DNA targeting by nucleosomes and chromatin

2021 ◽  
Vol 7 (11) ◽  
pp. eabd6030
Author(s):  
Isabel Strohkendl ◽  
Fatema A. Saifuddin ◽  
Bryan A. Gibson ◽  
Michael K. Rosen ◽  
Rick Russell ◽  
...  
Keyword(s):  
Histone Modifications ◽  
Dna Cleavage ◽  
Genome Engineering ◽  
Eukaryotic Cells ◽  
Loop Formation ◽  
Chromatin Compaction ◽  
Dna Targeting ◽  
Genomic Regions ◽  
Dna Bendability ◽  
Nucleosome Arrays

Genome engineering nucleases must access chromatinized DNA. Here, we investigate how AsCas12a cleaves DNA within human nucleosomes and phase-condensed nucleosome arrays. Using quantitative kinetics approaches, we show that dynamic nucleosome unwrapping regulates target accessibility to Cas12a and determines the extent to which both steps of binding—PAM recognition and R-loop formation—are inhibited by the nucleosome. Relaxing DNA wrapping within the nucleosome by reducing DNA bendability, adding histone modifications, or introducing target-proximal dCas9 enhances DNA cleavage rates over 10-fold. Unexpectedly, Cas12a readily cleaves internucleosomal linker DNA within chromatin-like, phase-separated nucleosome arrays. DNA targeting is reduced only ~5-fold due to neighboring nucleosomes and chromatin compaction. This work explains the observation that on-target cleavage within nucleosomes occurs less often than off-target cleavage within nucleosome-depleted genomic regions in cells. We conclude that nucleosome unwrapping regulates accessibility to CRISPR-Cas nucleases and propose that increasing nucleosome breathing dynamics will improve DNA targeting in eukaryotic cells.

scholarly journals Analysis of the chromosomal clustering of Fusarium-responsive wheat genes uncovers new players in the defence against head blight disease

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Alexandre Perochon ◽  
Harriet R. Benbow ◽  
Katarzyna Ślęczka-Brady ◽  
Keshav B. Malla ◽  
Fiona M. Doohan
Keyword(s):  
Map Kinases ◽  
Gene Families ◽  
Gene Clusters ◽  
Head Blight ◽  
Orphan Gene ◽  
Metabolic Genes ◽  
Eukaryotic Gene ◽  
Blight Disease ◽  
Genomic Regions ◽  
Eukaryotic Genomes

AbstractThere is increasing evidence that some functionally related, co-expressed genes cluster within eukaryotic genomes. We present a novel pipeline that delineates such eukaryotic gene clusters. Using this tool for bread wheat, we uncovered 44 clusters of genes that are responsive to the fungal pathogen Fusarium graminearum. As expected, these Fusarium-responsive gene clusters (FRGCs) included metabolic gene clusters, many of which are associated with disease resistance, but hitherto not described for wheat. However, the majority of the FRGCs are non-metabolic, many of which contain clusters of paralogues, including those implicated in plant disease responses, such as glutathione transferases, MAP kinases, and germin-like proteins. 20 of the FRGCs encode nonhomologous, non-metabolic genes (including defence-related genes). One of these clusters includes the characterised Fusarium resistance orphan gene, TaFROG. Eight of the FRGCs map within 6 FHB resistance loci. One small QTL on chromosome 7D (4.7 Mb) encodes eight Fusarium-responsive genes, five of which are within a FRGC. This study provides a new tool to identify genomic regions enriched in genes responsive to specific traits of interest and applied herein it highlighted gene families, genetic loci and biological pathways of importance in the response of wheat to disease.

scholarly journals Development of a loop-mediated isothermal amplification assay for the detection of Tilletia controversa based on genome comparison

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Somayyeh Sedaghatjoo ◽  
Monika K. Forster ◽  
Ludwig Niessen ◽  
Petr Karlovsky ◽  
Berta Killermann ◽  
...  
Keyword(s):  
Test Performance ◽  
Genomic Dna ◽  
Lamp Assay ◽  
Fungal Species ◽  
Isothermal Amplification ◽  
Genome Comparison ◽  
Performance Study ◽  
Loop Mediated Isothermal Amplification ◽  
Tilletia Controversa ◽  
Genomic Regions

AbstractTilletia controversa causing dwarf bunt of wheat is a quarantine pathogen in several countries. Therefore, its specific detection is of great phytosanitary importance. Genomic regions routinely used for phylogenetic inferences lack suitable polymorphisms for the development of species-specific markers. We therefore compared 21 genomes of six Tilletia species to identify DNA regions that were unique and conserved in all T. controversa isolates and had no or limited homology to other Tilletia species. A loop-mediated isothermal amplification (LAMP) assay for T. controversa was developed based on one of these DNA regions. The specificity of the assay was verified using 223 fungal samples comprising 43 fungal species including 11 Tilletia species, in particular 39 specimens of T. controversa, 92 of T. caries and 40 of T. laevis, respectively. The assay specifically amplified genomic DNA of T. controversa from pure cultures and teliospores. Only Tilletia trabutii generated false positive signals. The detection limit of the LAMP assay was 5 pg of genomic DNA per reaction. A test performance study that included five laboratories in Germany resulted in 100% sensitivity and 97.7% specificity of the assay. Genomic regions, specific to common bunt (Tilletia caries and Tilletia laevis together) are also provided.

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