scholarly journals De Novo Transcriptomic Characterization Enables Novel Microsatellite Identification and Marker Development in Betta splendens

Life ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 803
Author(s):  
Huapu Chen ◽  
Xiaomeng Li ◽  
Yaorong Wang ◽  
Chunhua Zhu ◽  
Hai Huang ◽  
...  
Keyword(s):  
Ssr Markers ◽  
De Novo ◽  
Habitat Degradation ◽  
Betta Splendens ◽  
Orthologous Group ◽  
Functional Genes ◽  
Genetics Research ◽  
Ssr Loci ◽  
Paired End Sequencing ◽  
Eukaryotic Orthologous Group

The wild populations of the commercially valuable ornamental fish species, Betta splendens, and its germplasm resources have long been threatened by habitat degradation and contamination with artificially bred fish. Because of the lack of effective marker resources, population genetics research projects are severely hampered. To generate genetic data for developing polymorphic simple sequence repeat (SSR) markers and identifying functional genes, transcriptomic analysis was performed. Illumina paired-end sequencing yielded 105,505,486 clean reads, which were then de novo assembled into 69,836 unigenes. Of these, 35,751 were annotated in the non-redundant, EuKaryotic Orthologous Group, Swiss-Prot, Kyoto Encyclopedia of Genes and Genomes and Gene Ontology databases. A total of 12,751 SSR loci were identified from the transcripts and 7970 primer pairs were designed. One hundred primer pairs were randomly selected for PCR validation and 53 successfully generated target amplification products. Further validation demonstrated that 36% (n = 19) of the 53 amplified loci were polymorphic. These data could not only enrich the genetic information for the identification of functional genes but also effectively facilitate the development of SSR markers. Such knowledge would accelerate further studies on the genetic variation and evolution, comparative genomics, linkage mapping and molecular breeding in B. splendens.

scholarly journals Development of polymorphic EST-SSR markers and characterization of the autotetraploid genome of sainfoin (Onobrychis viciifolia)

PeerJ ◽  
2019 ◽  
Vol 7 ◽  
pp. e6542 ◽  
Author(s):  
Shuheng Shen ◽  
Xutian Chai ◽  
Qiang Zhou ◽  
Dong Luo ◽  
Yanrong Wang ◽  
...  
Keyword(s):  
Ssr Markers ◽  
De Novo ◽  
Genetic Studies ◽  
Onobrychis Viciifolia ◽  
Ssr Loci ◽  
Wild Accessions ◽  
High Level ◽  
Genetic Structures ◽  
Paired End Sequencing

Background Sainfoin (Onobrychis viciifolia) is a highly nutritious, tannin-containing, and tetraploid forage legume. Due to the lack of detailed transcriptomic and genomic information on this species, genetic and breeding projects for sainfoin improvement have been significantly hindered. Methods In this study, a total of 24,630,711 clean reads were generated from 14 different sainfoin tissues using Illumina paired-end sequencing technology and deposited in the NCBI SRA database (SRX3763386). From these clean reads, 77,764 unigene sequences were obtained and 6,752 EST-SSRs were identified using de novo assembly. A total of 2,469 primer pairs were designed, and 200 primer pairs were randomly selected to analyze the polymorphism in five sainfoin wild accessions. Results Further analysis of 40 sainfoin individuals from the five wild populations using 61 EST-SSR loci showed that the number of alleles per locus ranged from 4 to 15, and the expected heterozygosity varied from 0.55 to 0.91. Additionally, by counting the EST-SSR band number and sequencing the three or four bands in one sainfoin individual, sainfoin was confirmed to be autotetraploid. This finding provides a high level of information about this plant. Discussion Through this study, 61 EST-SSR markers were successfully developed and shown to be useful for genetic studies and investigations of population genetic structures and variabilities among different sainfoin accessions.

Mining, characterization and application of transcriptome-based SSR markers in Chinese jiaotou

2018 ◽  
Vol 16 (4) ◽  
pp. 306-314
Author(s):  
Chan Liu ◽  
Qing Tang ◽  
Chaohua Cheng ◽  
Ying Xu ◽  
Zemao Yang ◽  
...  
Keyword(s):  
Ssr Markers ◽  
Large Scale ◽  
Trinucleotide Repeat ◽  
De Novo ◽  
Local Varieties ◽  
Genetic Studies ◽  
Oxidation Reduction ◽  
Ssr Loci ◽  
Wild Accessions ◽  
Expressed Sequence

AbstractChinese jiaotou is an economically important crop that is widely cultivated in East Asia. The lack of simple sequence repeat (SSR) markers has been a major obstacle for genetic studies of this crop. In the present study, SSR markers were developed for Chinese jiaotou on a large scale, based on the crop's transcriptome assembledde novoby a previous study. A search for SSR loci in the transcriptome's expressed sequence tags (ESTs) revealed 2157 SSRs, of which primer pairs could be developed for 1494. Among these resulting SSRs, trinucleotide repeat motifs were the most abundant type, with GAA/TTC motifs occurring most frequently. Analysing the annotated function of SSR-containing ESTs revealed that they enriched into the GO categories involved in transcription regulation, oxidation–reduction, transport, etc. The quality and transferability of these markers were also assessed using 100 randomly selected EST–SSRs, and the result showed that these markers were of good quality and possessed high cross-species transferability. In addition, the developed SSR markers were used to analyse the genetic diversity of 19 cultivated and four wild accessions, resulting in three distinct groups, cluster I, II and III. Interestingly, all four wild accessions were assigned to cluster III, and two local varieties from northern Hunan, China, were closely related to the wild genotypes. These results provide new insights into the origin of Chinese jiaotou. The EST–SSRs developed herein represent the first large-scale development of SSR markers in Chinese jiaotou, and they can be widely used for genetic studies of the crop.

scholarly journals Development of polymorphic EST-SSR markers and characterization of the autotetraploid in sainfoin (Onobrychis viciifolia)

2018 ◽  
Author(s):  
Shuheng Shen ◽  
Xutian Chai ◽  
Dong Luo ◽  
Yanrong Wang ◽  
Zhipeng Liu
Keyword(s):  
Ssr Markers ◽  
Genetic Studies ◽  
Onobrychis Viciifolia ◽  
Ssr Loci ◽  
Wild Accessions ◽  
High Level ◽  
Genetic Structures ◽  
Paired End Sequencing ◽  
Band Number

Background: Sainfoin (Onobrychis viciifolia) is a highly nutritious, tannin-containing, and tetraploid forage legume. Due to the lack of detailed transcriptomic and genomic information on this species, genetic and breeding projects for sainfoin improvement have been significantly hindered. Methods: In this study, a total of 24,630,711 clean reads were generated from 14 different sainfoin tissues using Illumina paired-end sequencing technology and deposited in the NCBI SRA database (SRX3763386). From these clean reads, 77,764 unigene sequences were obtained and 6,752 EST-SSRs were identified using denovo assembly. A total of 2,469 primer pairs were designed, and 200 primer pairs were randomly selected to analyze the polymorphism in five sainfoin wild accessions. Results: Further analysis of 40 sainfoin individuals from the five wild populations using 61 EST-SSR loci showed that the number of alleles per locus ranged from 4 to 15, and the expected heterozygosity varied from 0.55 to 0.91. Additionally, by counting the EST-SSR band number and sequencing the three or four bands in one sainfoin individual, sainfoin was confirmed to be autotetraploid. This finding provides a high level of information about this plant. Discussion: Through this study, 61 EST-SSR markers were successfully developed and shown to be useful for genetic studies and investigations of population genetic structures and variabilities among different sainfoin accessions.

scholarly journals Transcriptome Sequencing of Different Avocado Ecotypes: de novo Transcriptome Assembly, Annotation, Identification and Validation of EST-SSR Markers

Forests ◽  
2019 ◽  
Vol 10 (5) ◽  
pp. 411 ◽  
Author(s):  
Yu Ge ◽  
Lin Tan ◽  
Bin Wu ◽  
Tao Wang ◽  
Teng Zhang ◽  
...  
Keyword(s):  
Ssr Markers ◽  
High Throughput Sequencing ◽  
Expressed Sequence Tag ◽  
De Novo ◽  
Average Length ◽  
Transcriptome Assembly ◽  
Persea Americana ◽  
Illumina Hiseq ◽  
Ssr Loci ◽  
Ssr Development

Avocado (Persea americana Mill.) could be considered as an important tropical and subtropical woody oil crop with high economic and nutritional value. Despite the importance of this species, genomic information is currently unavailable for avocado and closely related congeners. In this study, we generated more than 216 million clean reads from different avocado ecotypes using Illumina HiSeq high-throughput sequencing technology. The high-quality reads were assembled into 154,310 unigenes with an average length of 922 bp. A total of 55,558 simple sequence repeat (SSR) loci detected among the 43,270 SSR-containing unigene sequences were used to develop 74,580 expressed sequence tag (EST)-SSR markers. From these markers, a subset of 100 EST-SSR markers was randomly chosen to identify polymorphic EST-SSR markers in 28 avocado accessions. Sixteen EST-SSR markers with moderate to high polymorphism levels were detected, with polymorphism information contents ranging from 0.33 to 0.84 and averaging 0.63. These 16 polymorphic EST-SSRs could clearly and effectively distinguish the 28 avocado accessions. In summary, our study is the first presentation of transcriptome data of different avocado ecotypes and comprehensive study on the development and analysis of a set of EST-SSR markers in avocado. The application of next-generation sequencing techniques for SSR development is a potentially powerful tool for genetic studies.

scholarly journals Development of polymorphic EST-SSR markers and characterization of the autotetraploid in sainfoin (Onobrychis viciifolia)

2018 ◽  
Author(s):  
Shuheng Shen ◽  
Xutian Chai ◽  
Dong Luo ◽  
Yanrong Wang ◽  
Zhipeng Liu
Keyword(s):  
Ssr Markers ◽  
Genetic Studies ◽  
Onobrychis Viciifolia ◽  
Ssr Loci ◽  
Wild Accessions ◽  
High Level ◽  
Genetic Structures ◽  
Paired End Sequencing ◽  
Band Number

Background: Sainfoin (Onobrychis viciifolia) is a highly nutritious, tannin-containing, and tetraploid forage legume. Due to the lack of detailed transcriptomic and genomic information on this species, genetic and breeding projects for sainfoin improvement have been significantly hindered. Methods: In this study, a total of 24,630,711 clean reads were generated from 14 different sainfoin tissues using Illumina paired-end sequencing technology and deposited in the NCBI SRA database (SRX3763386). From these clean reads, 77,764 unigene sequences were obtained and 6,752 EST-SSRs were identified using denovo assembly. A total of 2,469 primer pairs were designed, and 200 primer pairs were randomly selected to analyze the polymorphism in five sainfoin wild accessions. Results: Further analysis of 40 sainfoin individuals from the five wild populations using 61 EST-SSR loci showed that the number of alleles per locus ranged from 4 to 15, and the expected heterozygosity varied from 0.55 to 0.91. Additionally, by counting the EST-SSR band number and sequencing the three or four bands in one sainfoin individual, sainfoin was confirmed to be autotetraploid. This finding provides a high level of information about this plant. Discussion: Through this study, 61 EST-SSR markers were successfully developed and shown to be useful for genetic studies and investigations of population genetic structures and variabilities among different sainfoin accessions.

scholarly journals De novo transcriptomic analysis and identification of EST-SSR markers in Stephanandra incisa

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Cuiping Zhang ◽  
Zhonglan Wu ◽  
Xinqiang Jiang ◽  
Wei Li ◽  
Yizeng Lu ◽  
...  
Keyword(s):  
Ssr Markers ◽  
De Novo ◽  
Average Length ◽  
Resource Assessment ◽  
Sequence Information ◽  
Average Value ◽  
Information Index ◽  
Ssr Loci ◽  
Level Cluster ◽  
Polymorphism Level

AbstractStephanandra incisa is a wild-type shrub with beautiful leaves and white flowers and is commonly used as a garden decoration accessory. However, the limited availability of genomic data of S. incisa has restricted its breeding process. Here, we identified EST-SSR markers using de novo transcriptome sequencing. In this study, a transcriptome database containing 35,251 unigenes, having an average length of 985 bp, was obtained from S. incisa. From these unigene sequences, we identified 5,555 EST-SSRs, with a distribution density of one SSR per 1.60 kb. Dinucleotides (52.96%) were the most detected SSRs, followed by trinucleotides (34.64%). From the EST-SSR loci, we randomly selected 100 sites for designing primer and used the DNA of 60 samples to verify the polymorphism. The average value of the effective number of alleles (Ne), Shannon’s information index (I), and expective heterozygosity (He) was 1.969, 0.728, and 0.434, respectively. The polymorphism information content (PIC) value was in the range of 0.108 to 0.669, averaging 0.406, which represented a middle polymorphism level. Cluster analysis of S. incisa were also performed based on the obtained EST-SSR data in our work. As shown by structure analysis, 60 individuals could be classified into two groups. Thus, the identification of these novel EST-SSR markers provided valuable sequence information for analyzing the population structure, genetic diversity, and genetic resource assessment of S. incisa and other related species.

scholarly journals Characterization of metabolic responses, genetic variations, and microsatellite instability in ammonia-stressed CHO cells grown in fed-batch cultures

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Dylan G. Chitwood ◽  
Qinghua Wang ◽  
Kathryn Elliott ◽  
Aiyana Bullock ◽  
Dwon Jordana ◽  
...  
Keyword(s):  
Microsatellite Instability ◽  
Microsatellite Loci ◽  
Cho Cells ◽  
Genome Stability ◽  
De Novo ◽  
Genome Instability ◽  
Chinese Hamster ◽  
Functional Genes ◽  
Nucleotide Polymorphisms ◽  
De Novo Mutations

Abstract Background As bioprocess intensification has increased over the last 30 years, yields from mammalian cell processes have increased from 10’s of milligrams to over 10’s of grams per liter. Most of these gains in productivity can be attributed to increasing cell densities within bioreactors. As such, strategies have been developed to minimize accumulation of metabolic wastes, such as lactate and ammonia. Unfortunately, neither cell growth nor biopharmaceutical production can occur without some waste metabolite accumulation. Inevitably, metabolic waste accumulation leads to decline and termination of the culture. While it is understood that the accumulation of these unwanted compounds imparts a suboptimal culture environment, little is known about the genotoxic properties of these compounds that may lead to global genome instability. In this study, we examined the effects of high and moderate extracellular ammonia on the physiology and genomic integrity of Chinese hamster ovary (CHO) cells. Results Through whole genome sequencing, we discovered 2394 variant sites within functional genes comprised of both single nucleotide polymorphisms and insertion/deletion mutations as a result of ammonia stress with high or moderate impact on functional genes. Furthermore, several of these de novo mutations were found in genes whose functions are to maintain genome stability, such as Tp53, Tnfsf11, Brca1, as well as Nfkb1. Furthermore, we characterized microsatellite content of the cultures using the CriGri-PICR Chinese hamster genome assembly and discovered an abundance of microsatellite loci that are not replicated faithfully in the ammonia-stressed cultures. Unfaithful replication of these loci is a signature of microsatellite instability. With rigorous filtering, we found 124 candidate microsatellite loci that may be suitable for further investigation to determine whether these loci may be reliable biomarkers to predict genome instability in CHO cultures. Conclusion This study advances our knowledge with regards to the effects of ammonia accumulation on CHO cell culture performance by identifying ammonia-sensitive genes linked to genome stability and lays the foundation for the development of a new diagnostic tool for assessing genome stability.

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