PCR-based evaluation of sequence specificity of DNA fragmentation by ultrasound

Гарафутдинов, Р.Р.; Galimova, A. A.; Сахабутдинова, А. Р.; Чемерис, А. В.

doi:10.1134/s0026893316020059

Cited by 6 publications

(3 citation statements)

References 18 publications

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“…This is usually achieved using ultrasonicators such as Bioruptor or Covaris. Some studies have reported that this process does not occur randomly (Garafutdinov, Galimova, Sakhabutdinova, & Chemeris, ; Poptsova et al, ), but depends on the nucleotide sequences. The effect of such nonrandom fragmentation has not been assessed in the context of diet inference from eDNA, but might also introduce distortion, and thus, processing all samples with the same shearing mechanism and protocol is advisable.…”

Section: Factors Distorting Diet Assessmentsmentioning

confidence: 99%

Promises and pitfalls of using high‐throughput sequencing for diet analysis

Alberdi

Aizpurua

Bohmann

et al. 2018

Molecular Ecology Resources

173

195

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The application of high‐throughput sequencing‐based approaches to DNA extracted from environmental samples such as gut contents and faeces has become a popular tool for studying dietary habits of animals. Due to the high resolution and prey detection capacity they provide, both metabarcoding and shotgun sequencing are increasingly used to address ecological questions grounded in dietary relationships. Despite their great promise in this context, recent research has unveiled how a wealth of biological (related to the study system) and technical (related to the methodology) factors can distort the signal of taxonomic composition and diversity. Here, we review these studies in the light of high‐throughput sequencing‐based assessment of trophic interactions. We address how the study design can account for distortion factors, and how acknowledging limitations and biases inherent to sequencing‐based diet analyses are essential for obtaining reliable results, thus drawing appropriate conclusions. Furthermore, we suggest strategies to minimize the effect of distortion factors, measures to increase reproducibility, replicability and comparability of studies, and options to scale up DNA sequencing‐based diet analyses. In doing so, we aim to aid end‐users in designing reliable diet studies by informing them about the complexity and limitations of DNA sequencing‐based diet analyses, and encourage researchers to create and improve tools that will eventually drive this field to its maturity.

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Section: Factors Distorting Diet Assessmentsmentioning

confidence: 99%

Promises and pitfalls of using high‐throughput sequencing for diet analysis

Alberdi

Aizpurua

Bohmann

et al. 2018

Molecular Ecology Resources

173

195

View full text Add to dashboard Cite

show abstract

“…A possible explanation is that exon 1 was deleted from the S ‐locus (causing CYP T loss of function, hence homostyly) and translocated to a highly repetitive genomic region. The translocation could have allowed targeted amplification and subsequent Sanger sequencing using exon‐1‐specific PCR primers, while preventing exon‐1 detection via next generation sequencing due to biases arising, for example, during genomic DNA sonication used to produce short DNA fragments prior to short‐read library preparation (Garafutdinov et al., 2016 ; Jennings et al., 2017 ; Poptsova et al., 2014 ). Notably, a few low‐quality sequencing reads did map to CYP T exon 1, suggesting this exon is indeed present in the genome of these homostyles but was not successfully sequenced using short‐read sequencing methodology.…”

Section: Discussionmentioning

confidence: 99%

Genomic analyses elucidate S‐locus evolution in response to intra‐specific losses of distyly in Primula vulgaris

Mora‐Carrera,

Stubbs,

Potente

et al. 2024

Ecology and Evolution

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Distyly, a floral dimorphism that promotes outcrossing, is controlled by a hemizygous genomic region known as the S‐locus. Disruptions of genes within the S‐locus are responsible for the loss of distyly and the emergence of homostyly, a floral monomorphism that favors selfing. Using whole‐genome resequencing data of distylous and homostylous individuals from populations of Primula vulgaris and leveraging high‐quality reference genomes of Primula we tested, for the first time, predictions about the evolutionary consequences of transitions to selfing on S‐genes. Our results reveal a previously undetected structural rearrangement in CYPᵀ associated with the shift to homostyly and confirm previously reported, homostyle‐specific, loss‐of‐function mutations in the exons of the S‐gene CYPᵀ. We also discovered that the promoter and intronic regions of CYPᵀ in distylous and homostylous individuals are conserved, suggesting that down‐regulation of CYPᵀ via mutations in its promoter and intronic regions is not a cause of the shift to homostyly. Furthermore, we found that hemizygosity is associated with reduced genetic diversity in S‐genes compared with their paralogs outside the S‐locus. Additionally, the shift to homostyly lowers genetic diversity in both the S‐genes and their paralogs, as expected in primarily selfing plants. Finally, we tested, for the first time, long‐standing theoretical models of changes in S‐locus genotypes during early stages of the transition to homostyly, supporting the assumption that two copies of the S‐locus might reduce homostyle fitness.

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“…A possible explanation is that exon 1 was deleted from the S-locus (causingCYP T loss of function, hence homostyly) and translocated to a highly repetitive genomic region. The translocation could have allowed targeted amplification and subsequent Sanger sequencing using exon-1-specific PCR primers, while preventing exon-1 detection via next generation sequencing due to biases arising, for example, during genomic DNA sonication used to produce short DNA fragments prior to short-read library preparation (Poptsova et al , 2014;Garafutdinov et al , 2016;Jennings et al , 2017). Notably, a few low-quality sequencing reads did map toCYP T exon 1, suggesting this exon is indeed present in the genome of these homostyles but was not successfully sequenced using short-read sequencing methodology.…”

Section: Genetic Basis Of Transitions From Distyly To Homostyly In Pr...mentioning

confidence: 99%

Genomic analyses elucidate S-locus evolution in response to intra-specific losses of distyly Primula vulgaris

Mora‐Carrera

Stubbs

Potente

et al. 2023

Preprint

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Distyly, a floral dimorphism that promotes outcrossing, is controlled by a hemizygous genomic region known as the S-locus. Disruptions of genes within the S-locus are responsible for the loss of distyly and the emergence of homostyly, a floral monomorphism that favors selfing. Using whole genome resequencing data of distylous and homostylous individuals from populations of Primula vulgaris and leveraging high-quality reference genomes of Primula we tested, for the first time, predictions about the evolutionary consequences of transitions to selfing on S-locus genes. Our results confirm the presence of previously reported homostyle-specific, loss-of-function mutations in the exons of the S-locus gene CYPᵀ, while also revealing a previously undetected structural rearrangement in CYPᵀ associated with the shift to homostyly. Additionally, we discovered that the promoter region of CYPᵀ in distylous and homostylous individuals is identical, suggesting that down-regulation of CYPᵀ via mutations in its promoter region is not a cause of shift to homostyly. Furthermore, we found that hemizygosity leads to reduced genetic diversity and less efficient purifying selection in S-locus genes compared to genes outside the S-locus, and that the shift to homostyly further lowers genetic diversity, as expected for mating-system shifts. Finally, we tested, for the first time, long-standing theoretical models of changes in S-locus genotypes during early stages of the transition to homostyly, supporting the assumption that two (diploid) copies of the S-locus might reduce homostyle viability.

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PCR-based evaluation of sequence specificity of DNA fragmentation by ultrasound

Cited by 6 publications

References 18 publications

Promises and pitfalls of using high‐throughput sequencing for diet analysis

Promises and pitfalls of using high‐throughput sequencing for diet analysis

Genomic analyses elucidate S‐locus evolution in response to intra‐specific losses of distyly in Primula vulgaris

Genomic analyses elucidate S-locus evolution in response to intra-specific losses of distyly Primula vulgaris

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