Genomic Resources of Broomcorn Millet: Demonstration and Application of a High-throughput BAC Mapping Pipeline

AbstractBackgroundWith high-efficient water-use and drought tolerance, broomcorn millet has emerged as candidate for food security. To promote its research process for molecular breeding and functional research, a comprehensive genome resources is of great important.ResultsHerein, we constructed the first BAC library for broomcorn millet, generated BAC end sequences and integrated BAC clones into genome by a novel pipeline for BAC end profiling depending on clone-array pooled shotgun sequencing strategy and Illumina sequencing technology. The BAC library is consisted of 76,023 clones with average insert length of 123.48 Kb, about 9.9x coverage of 850 Mb genome. Then, 8262 of 9216 clones were mapped on broomcorn millet cultivar longmi4 genome using our pipeline. Furthermore, we also extracted and assembled unmapped reads against longmi4 genome. A total of 135 deletion sequences, 64 specific sequences and some sample contamination sequences were identified.ConclusionsBAC clones in this library can be browsed and obtained from our website (http://flyinguineapig.com/gresource/JBrowse-1.16.5/index.html). This pipeline designed for BAC end profiling can greatly reduce the cost of acquiring BAC end sequences and shorten the period of the experiment compared with the Sanger sequencing method. These high-quality BAC clones related with genome in this study provide a useful and convinient genomic resource for genome gap filling, complex segment sequencing, FISH, functional research, and genetic engineering.

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Genomic resources of broomcorn millet: demonstration and application of a high-throughput BAC mapping pipeline

BMC Genomic Data ◽

10.1186/s12863-021-01003-z ◽

2021 ◽

Vol 22 (1) ◽

Author(s):

Wei Xu ◽

Mengjie Liang ◽

Xue Yang ◽

Hao Wang ◽

Meizhong Luo

Keyword(s):

Bac Library ◽

Research Process ◽

High Quality ◽

Bac Clones ◽

Broomcorn Millet ◽

High Efficient ◽

Sequencing Strategy ◽

End Sequences ◽

Genomic Resource ◽

Efficient Water Use

Abstract Background With high-efficient water-use and drought tolerance, broomcorn millet has emerged as a candidate for food security. To promote its research process for molecular breeding and functional research, a comprehensive genome resource is of great importance. Results Herein, we constructed a BAC library for broomcorn millet, generated BAC end sequences based on the clone-array pooled shotgun sequencing strategy and Illumina sequencing technology, and integrated BAC clones into genome by a novel pipeline for BAC end profiling. The BAC library consisted of 76,023 clones with an average insert length of 123.48 Kb, covering about 9.9-fold of the 850 Mb genome. Of 9216 clones tested using our pipeline, 8262 clones were mapped on the broomcorn millet cultivar longmi4 genome. These mapped clones covered 308 of the 829 gaps left by the genome. To our knowledge, this is the only BAC resource for broomcorn millet. Conclusions We constructed a high-quality BAC libraray for broomcorn millet and designed a novel pipeline for BAC end profiling. BAC clones can be browsed and obtained from our website (http://eightstarsbio.com/gresource/JBrowse-1.16.5/index.html). The high-quality BAC clones mapped on genome in this study will provide a powerful genomic resource for genome gap filling, complex segment sequencing, FISH, functional research and genetic engineering of broomcorn millet.

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Genomic Resources of Broomcorn Millet: Demonstration and Application of a High-throughput BAC Mapping Pipeline

10.21203/rs.3.rs-536711/v1 ◽

2021 ◽

Author(s):

Wei Xu ◽

Mengjie Liang ◽

Xue Yang ◽

Hao Wang ◽

Meizhong Luo

Keyword(s):

Bac Library ◽

Research Process ◽

High Quality ◽

Bac Clones ◽

Broomcorn Millet ◽

High Efficient ◽

Sequencing Strategy ◽

End Sequences ◽

Genomic Resource ◽

Efficient Water Use

Abstract Background: With high-efficient water-use and drought tolerance, broomcorn millet has emerged as a candidate for food security. To promote its research process for molecular breeding and functional research, a comprehensive genome resource is of great importance. Results: Herein, we constructed a BAC library for broomcorn millet, generated BAC end sequences based on the clone-array pooled shotgun sequencing strategy and Illumina sequencing technology, and integrated BAC clones into genome by a novel pipeline for BAC end profiling. The BAC library is consisted of 76,023 clones with an average insert length of 123.48 Kb, covering about 9.9-fold of the 850 Mb genome. Of 9,216 clones tested using our pipeline, 8,262 clones were mapped on the broomcorn millet cultivar longmi4 genome. These mapped clones covered 308 of the 829 gaps left by the genome. To our knowledge, this is the only BAC resource for broomcorn millet.Conclusions: We constructed a high-quality BAC libraray for broomcorn millet and designed a novel pipeline for BAC end profiling. BAC clones can be browsed and obtained from our website (http://eightstarsbio.com/gresource/JBrowse-1.16.5/index.html). The high-quality BAC clones mapped on genome in this study will provide a powerful genomic resource for genome gap filling, complex segment sequencing, FISH, functional research, and genetic engineering of broomcorn millet.

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A Short Sequence Splicing Method for Genome Assembly Using a Three-Dimensional Mixing-Pool of BAC Clones and High-throughput Technology

The Open Biotechnology Journal ◽

10.2174/1874070701509010210 ◽

2015 ◽

Vol 9 (1) ◽

pp. 210-215

Author(s):

Xiaojun Kang ◽

Cheng Yang ◽

Xuguang Zhao ◽

Weiwei Chen ◽

Sifa Zhang ◽

...

Keyword(s):

High Throughput ◽

High Throughput Sequencing ◽

Three Dimensional ◽

Whole Genome Sequence ◽

Sequencing Data ◽

Assembly Algorithm ◽

Bac Clones ◽

High Throughput Technology ◽

Sequencing Strategy ◽

The Cost

Current genome sequencing techniques are expensive, and it is still a major challenge to obtain an individual whole-genome sequence. To reduce the cost of sequencing, this paper introduced a high-throughput sequencing strategy using a three-dimensional mixing-pools based on the cube. Following the strategy, BAC clones were injected into each vertex of the cube, and sequencing of each plane provided information about multiple clones, thereby significantly reducing the cost of sequencing. In addition, Velvet was used to assemble the sequencing data. The scaffold generated from Velvet contained a number of contigs, which were orderless. Therefore, to address this problem, a scaffold assembly algorithm based on multi-way trees was used. The algorithm used a multi-way tree to build the framework of chromosomes, and subsequently, the frame was filled to complete the scaffold assembly. This algorithm alone outperformed Velvet in the assembling of a scaffold.

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Screening of apple genotypes with the columnar growth habit using control markers

BIO Web of Conferences ◽

10.1051/bioconf/20202503007 ◽

2020 ◽

Vol 25 ◽

pp. 03007

Author(s):

Natalia Saveleva ◽

Alexander Lyzhin ◽

Andrey Yushkov ◽

Alexander Zemisov ◽

Nadezhda Borzykh

Keyword(s):

Growth Habit ◽

Early Stage ◽

Research Process ◽

Commercial Production ◽

Juvenile Stage ◽

Juvenile Period ◽

Columnar Growth ◽

Apple Cultivars ◽

The Cost ◽

Modern Breeding

One of the approaches to intensify horticulture is to introduce cultivars with an unusual canopy into commercial production. Such plants can be columnar Apple trees. In modern breeding, there is a trend to create cultivars with a compact canopy. In such orchards, it is reduced to a minimum the cost for pruning, harvesting, and protection against pest and disease, which are the main expenses in apple orchards with a traditional canopy. The use of molecular markers linked to columnar growth habit allows us to identify a physiological sign at an early stage of growth: in the juvenile period. The assessment of apple cultivars and hybrids was carried out at the I. V. Michurin Federal Scientific Centre in 2015-2018. Four markers were used in the research: Mdo. chr 10.12, C18470-25831, 29f1, and jwlr to identify plants with the columnar growth habit gene (Co). The use of various DNA markers made it possible to establish that not all of them are well linked to the Co gene. In the research process, primers were identified for markers 29f1 and jwlr, which reliably allowed us to identify plants with columnar growth habit at the juvenile stage, which will significantly reduce the breeding process.

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Construction and characterization of bacterial artificial chromosome library of black-handed spider monkey (Ateles geoffroyi)

Genome ◽

10.1139/g03-122 ◽

2004 ◽

Vol 47 (2) ◽

pp. 239-245 ◽

Cited By ~ 4

Author(s):

Yaping Qian ◽

Li Jin ◽

Bing Su

Keyword(s):

Bacterial Artificial Chromosome ◽

Large Scale ◽

Bac Library ◽

Artificial Chromosome ◽

Spider Monkey ◽

Ateles Geoffroyi ◽

Comparative Genomic ◽

Average Insert Size ◽

Bac Clones ◽

Genomic Study

The large-insert genomic DNA library is a critical resource for genome-wide genetic dissection of target species. We constructed a high-redundancy bacterial artificial chromosome (BAC) library of a New World monkey species, the black-handed spider monkey (Ateles geoffroyi). A total of 193 152 BAC clones were generated in this library. The average insert size of the BAC clones was estimated to be 184.6 kb with the small inserts (50-100 kb) accounting for less than 3% and the non-recombinant clones only 1.2%. Assuming a similar genome size with humans, the spider monkey BAC library has about 11× genome coverage. In addition, by end sequencing of randomly selected BAC clones, we generated 367 sequence tags for the library. When blasted against human genome, they showed a good correlation between the number of hit clones and the size of the chromosomes, an indication of unbiased chromosomal distribution of the library. This black-handed spider monkey BAC library would serve as a valuable resource in comparative genomic study and large-scale genome sequencing of nonhuman primates.Key words: black-handed spider monkeys, Ateles geoffroyi, BAC library.

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One-Step Bark-Like Imitated Polypropylene (PP)/Polycarbonate (PC) Nanofibrous Meltblown Membrane for Efficient Particulate Matter Removal

Polymers ◽

10.3390/polym11081307 ◽

2019 ◽

Vol 11 (8) ◽

pp. 1307 ◽

Cited By ~ 4

Author(s):

Ting-Ting Li ◽

Xixi Cen ◽

Hai-Tao Ren ◽

Fei Sun ◽

Qi Lin ◽

...

Keyword(s):

Particulate Matter ◽

Fiber Diameter ◽

Average Fiber Diameter ◽

Efficient Removal ◽

Filtration Performance ◽

Innovative Method ◽

High Efficient ◽

One Step ◽

The Cost ◽

Pm Removal

A bark-like imitated polypr opylene (PP)/polycarbonate (PC) nanofibrous membrane was constructed by one-step meltblown technique for efficient particulate matter (PM) removal. The effects of PC content (0%, 1%, 3%, 5%, and 7%) on membrane thermal stability, microscopic characteristics, filtration performance, hydrophilicity, and water vapor transmission were investigated. The results demonstrated that using facile design of incompatibility and viscosity difference between PC and PP polymers decreases average fiber diameter, creating a bark-like groove appearance and increasing surface potential, making a new PP/PC membrane with high filtration performance. The resultant PP/PC membrane had finer average fiber diameter of 0.63 μm, which was nearly 89.41% lower than PP membranes (5.95 μm), and its quality factor (0.036 Pa−1) was nearly 2.12 times than that of PP membranes (0.017 Pa−1) with the die hole diameter of 0.5 mm. This fabrication technique of a special meltblown filter membrane saves the cost of die retrofitting and post-processing, which provides an innovative method for particulate efficient removal of high efficient filters.

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Use of a Mycobacterium tuberculosisH37Rv Bacterial Artificial Chromosome Library for Genome Mapping, Sequencing, and Comparative Genomics

Infection and Immunity ◽

10.1128/iai.66.5.2221-2229.1998 ◽

1998 ◽

Vol 66 (5) ◽

pp. 2221-2229 ◽

Cited By ~ 144

Author(s):

Roland Brosch ◽

Stephen V. Gordon ◽

Alain Billault ◽

Thierry Garnier ◽

Karin Eiglmeier ◽

...

Keyword(s):

Comparative Genomics ◽

Bacterial Artificial Chromosome ◽

Genome Mapping ◽

Sequence Data ◽

Bac Library ◽

Artificial Chromosome ◽

Excellent Correlation ◽

Sequencing Project ◽

Polysaccharide Biosynthesis ◽

Bac Clones

ABSTRACT The bacterial artificial chromosome (BAC) cloning system is capable of stably propagating large, complex DNA inserts in Escherichia coli. As part of the Mycobacterium tuberculosis H37Rv genome sequencing project, a BAC library was constructed in the pBeloBAC11 vector and used for genome mapping, confirmation of sequence assembly, and sequencing. The library contains about 5,000 BAC clones, with inserts ranging in size from 25 to 104 kb, representing theoretically a 70-fold coverage of the M. tuberculosisgenome (4.4 Mb). A total of 840 sequences from the T7 and SP6 termini of 420 BACs were determined and compared to those of a partial genomic database. These sequences showed excellent correlation between the estimated sizes and positions of the BAC clones and the sizes and positions of previously sequenced cosmids and the resulting contigs. Many BAC clones represent linking clones between sequenced cosmids, allowing full coverage of the H37Rv chromosome, and they are now being shotgun sequenced in the framework of the H37Rv sequencing project. Also, no chimeric, deleted, or rearranged BAC clones were detected, which was of major importance for the correct mapping and assembly of the H37Rv sequence. The minimal overlapping set contains 68 unique BAC clones and spans the whole H37Rv chromosome with the exception of a single gap of ∼150 kb. As a postgenomic application, the canonical BAC set was used in a comparative study to reveal chromosomal polymorphisms between M. tuberculosis, M. bovis, and M. bovis BCG Pasteur, and a novel 12.7-kb segment present in M. tuberculosis but absent from M. bovis and M. bovis BCG was characterized. This region contains a set of genes whose products show low similarity to proteins involved in polysaccharide biosynthesis. The H37Rv BAC library therefore provides us with a powerful tool both for the generation and confirmation of sequence data as well as for comparative genomics and other postgenomic applications. It represents a major resource for present and future M. tuberculosis research projects.

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PCR identification of durum wheat BAC clones containing genes coding for carotenoid biosynthesis enzymes and their chromosome localization

Genome ◽

10.1139/g04-033 ◽

2004 ◽

Vol 47 (5) ◽

pp. 911-917 ◽

Cited By ~ 32

Author(s):

A Cenci ◽

S Somma ◽

N Chantret ◽

J Dubcovsky ◽

A Blanco

Keyword(s):

Tetraploid Wheat ◽

Bac Library ◽

Pcr Amplification ◽

Carotenoid Biosynthesis ◽

Quality Trait ◽

Substitution Lines ◽

D Genome ◽

Pcr Screening ◽

Bac Clones ◽

Essential Components

Carotenoids are essential components in all plants. Their accumulation in wheat seed determines the endosperm colour, which is an important quality trait in wheat. In this study, we report the isolation of BAC clones containing genes coding for three different enzymes of the carotenoid biosynthesis pathway: phytoene synthase (PSY), phytoene desaturase (PDS), and ζ-carotene desaturase (ZDS). Primers were designed on the basis of wheat ESTs similar to the sequences of these three genes in other species, and used to screen a BAC library from Triticum turgidum var. durum (2n = 28, genomes AABB). Eight, six, and nine 384-well plates containing at least one positive clone were found for PSY, PDS, and ZDS, respectively. BACs selected for each of these genes were then divided in two groups corresponding to the A and B genomes of tetraploid wheat, based on differences in the length of the PCR amplification products, conformation-sensitive gel electrophoresis (CSGE), or cleavage amplification polymorphisms. Positive clones were then assigned to chromosomes using a set of D genome substitution lines in T. turgidum var. durum 'Langdon'. PSY clones were localized on chromosomes 5A and 5B, PDS on chromosomes 4A and 4B, and ZDS on chromosomes 2A and 2B. The strategies used for the PCR screening of large BAC libraries and for the differentiation of BAC clones from different genomes in a polyploid species are discussed.Key words: wheat, carotenoid biosynthesis, BAC.

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Construction of a hexaploid wheat (Triticum aestivumL.) bacterial artificial chromosome library for cloning genes for stripe rust resistance

Genome ◽

10.1139/g05-078 ◽

2005 ◽

Vol 48 (6) ◽

pp. 1028-1036 ◽

Cited By ~ 13

Author(s):

P Ling ◽

X M Chen

Keyword(s):

Triticum Aestivum ◽

Stripe Rust ◽

Hexaploid Wheat ◽

Rust Resistance ◽

Puccinia Striiformis ◽

Bac Library ◽

Artificial Chromosome ◽

Stripe Rust Resistance ◽

Insert Size ◽

Bac Clones

A hexaploid wheat (Triticum aestivum L.) bacterial artificial chromosome (BAC) library was constructed for cloning Yr5 and other genes conferring resistance to stripe rust (Puccinia striiformis f. sp. tritici). Intact nuclei from a Yr5 near-isogenic line were used to isolate high molecular weight DNA, which was partially cleaved with HindIII and cloned into pECBAC1 and pIndigoBAC-5 vectors. The wheat BAC library consisted of 422 400 clones arrayed in 1100 micro-titer plates (each plate with 384 wells). Random sampling of 300 BAC clones indicated an average insert size of 140 kb, with a size range from 25 to 365 kb. Ninety percent of the clones in the library had an insert size greater than 100 kb and fewer than 5% of the clones did not contain inserts. Based on an estimated genome size of 15 966 Mb for hexaploid wheat, the BAC library was estimated to have a total coverage of 3.58× wheat genome equivalents, giving approximately 96% probability of identifying a clone representing any given wheat DNA sequence. Twelve BAC clones containing an Yr5 locus-specific marker (Yr5STS7/8) were successfully selected by PCR screening of 3-dimensional BAC pools. The results demonstrated that the T. aestivum BAC library is a valuable genomic resource for positional cloning of Yr5. The library also should be useful in cloning other genes for stripe rust resistance and other traits of interest in hexaploid wheat.Key words: BAC library, BAC pools, hexaploid wheat, Puccinia striiformis f. sp. tritici, resistance gene, stripe rust, Triticum aestivum.

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Construction, Characterization, and Preliminary BAC-End Sequence Analysis of a Bacterial Artificial Chromosome Library of the Tea Plant (Camellia sinensis)

Journal of Biomedicine and Biotechnology ◽

10.1155/2011/476723 ◽

2011 ◽

Vol 2011 ◽

pp. 1-8 ◽

Cited By ~ 4

Author(s):

Jinke Lin ◽

Dave Kudrna ◽

Rod A. Wing

Keyword(s):

Camellia Sinensis ◽

Bac Library ◽

Gc Content ◽

Artificial Chromosome ◽

Tea Plant ◽

Average Insert Size ◽

Plant Genomics ◽

Insert Size ◽

Sequence Class ◽

End Sequences

We describe the construction and characterization of a publicly available BAC library for the tea plant,Camellia sinensis. Using modified methods, the library was constructed with the aim of developing public molecular resources to advance tea plant genomics research. The library consists of a total of 401,280 clones with an average insert size of 135 kb, providing an approximate coverage of 13.5 haploid genome equivalents. No empty vector clones were observed in a random sampling of 576 BAC clones. Further analysis of 182 BAC-end sequences from randomly selected clones revealed a GC content of 40.35% and low chloroplast and mitochondrial contamination. Repetitive sequence analyses indicated that LTR retrotransposons were the most predominant sequence class (86.93%–87.24%), followed by DNA retrotransposons (11.16%–11.69%). Additionally, we found 25 simple sequence repeats (SSRs) that could potentially be used as genetic markers.

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