Common Microsatellite Research Articles

A robust microsatellite instability detection model for unpaired colorectal cancer tissue samples.

Microsatellite instability (MSI) is a key biomarker for cancer immunotherapy and prognosis. Integration of MSI testing into a next-generation-sequencing (NGS) panel could save tissue sample, reduce turn-around time and cost, and provide MSI status and comprehensive genomic profiling in single test. We aimed to develop an MSI calling model to detect MSI status along with the NGS panel-based profiling test using tumor-only samples. From January 2019 to December 2020, a total of 174 colorectal cancer (CRC) patients were enrolled, including 31 MSI-high (MSI-H) and 143 microsatellite stability (MSS) cases. Among them, 56 paired tumor and normal samples (10 MSI-H and 46 MSS) were used for modeling, and another 118 tumor-only samples were used for validation. MSI polymerase chain reaction (MSI-PCR) was performed as the gold standard. A baseline was built for the selected microsatellite loci using the NGS data of 56 normal blood samples. An MSI detection model was constructed by analyzing the NGS data of tissue samples. The performance of the model was compared with the results of MSI-PCR. We first intersected the target genomic regions of the NGS panels used in this study to select common microsatellite loci. A total of 42 loci including 23 mononucleotide repeat sites and 19 longer repeat sites were candidates for modeling. As mononucleotide repeat sites are more sensitive and specific for detecting MSI status than sites with longer length motif and the mononucleotide repeat sites performed even better than the total sites, a model containing 23 mononucleotide repeat sites was constructed and named Colorectal Cancer Microsatellite Instability test (CRC-MSI). The model achieved 100% sensitivity and 100% specificity when compared with MSI-PCR in both training and validation sets. Furthermore, the CRC-MSI model was robust with the tumor content as low as 6%. In addition, 8 out of 10 MSI-H samples showed alternations in the four mismatch repair genes (MLH1, MSH2, MSH6, and PMS2). MSI status can be accurately determined along the targeted NGS panels using only tumor samples. The performance of mononucleotide repeat sites surpasses loci with longer repeat motif in MSI calling.

AutomAted RepeaT Identifier (AARTI): A tool to identify common, polymorphic, and unique microsatellites

Here we are presenting an automated computational pipeline used to mine 5976 mitochondrial genomes to identify common, polymorphic, and unique microsatellites also known as simple sequence repeats (SSRs). Microsatellites are repetitive motifs of 1–6 bases in a DNA sequence. Due to their abundance and highly polymorphic nature, microsatellites have become one of the widely used molecular/genetic markers valuable for many studies including gene tagging, genetic diversity, and species identification. Several computational tools dedicated to mine and categorize microsatellites in nucleotide sequences were developed; however, there is no tool which can identify unique, common and polymorphic microsatellites between each pair of nucleotide sequences. To explore such microsatellites, we have developed a fully automated computational pipeline named AutomAted RepeaT Identifier (AARTI). The AARTI is the only tool till date, which identifies common, polymorphic, and unique microsatellites between each pair of nucleotide sequences. The computational pipeline was constructed using the PERL programming language and the web server for the pipeline was developed with the help of PHP, HTML, CSS, and JavaScript. It was successfully tested to reproduce the results of previous study on 7 mitochondrial genomes of genus Orthotrichum. Moreover, the pipeline was also applied on 5846 (Metazoa) and 130 (Viridiplantae) mitochondrial genomes. The AARTI is freely available at https://lms.snu.edu.in/aarti/ and will certainly accelerate the studies of length variation in microsatellites between species. Additionally, it will be useful in comparative genomic studies, for the computational characterization of microsatellites, and has the potential to be a routine genome analysis pipeline for mitochondrial genomes.

Concept and Design of Martian Far-IR ORE Spectrometer (MIRORES)

Sulfide ores are a major source of noble (Au, Ag, and Pt) and base (Cu, Pb, Zn, Sn, Co, Ni, etc.) metals and will, therefore, be vital for the self-sustainment of future Mars colonies. Martian meteorites are rich in sulfides, which is reflected in recent findings for surface Martian rocks analyzed by the Spirit and Curiosity rovers. However, the only high-resolution (18 m/pixel) infrared (IR) spectrometer orbiting Mars, the Compact Reconnaissance Imaging Spectrometer for Mars (CRISM), onboard the Mars Reconnaissance Orbiter (MRO), is not well-suited for detecting sulfides on the Martian surface. Spectral interference with silicates impedes sulfide detection in the 0.4–3.9 μm CRISM range. In contrast, at least three common hydrothermal sulfides on Earth and Mars (pyrite, chalcopyrite, marcasite) have prominent absorption peaks in a narrow far-IR (FIR) wavelength range of 23–28 μm. Identifying the global distribution and chemical composition of sulfide ore deposits would help in choosing useful targets for future Mars exploration missions. Therefore, we have designed a new instrument suitable for measuring sulfides in the FIR range called the Martian far-IR Ore Spectrometer (MIRORES). MIRORES will measure radiation in six narrow bands (~0.3 µm in width), including three bands centered on the sulfide absorption bands (23.2, 24.3 and 27.6 µm), two reference bands (21.5 and 26.1) and one band for clinopyroxene interference (29.0 µm). Focusing on sulfides only will make it possible to adapt the instrument size (32 × 32 × 42 cm) and mass (<10 kg) to common microsatellite requirements. The biggest challenges related to this design are: (1) the small field of view conditioned by the high resolution required for such a study (<20 m/pixel), which, in limited space, can only be achieved by the use of the Cassegrain optical system; and (2) a relatively stable measurement temperature to maintain radiometric accuracy and enable precise calibration.

Gyro-stellar inertial attitude estimation for satellite with high motion rate

AbstractFor a common micro-satellite, orbiting in a circular sun-synchronous orbit (SSO) at an altitude between 500 and 600km, the satellite attitude during off-nadir imaging and staring-imaging operations can be up to ±45 degree on roll and pitch angles. During these off-nadir pointing for both multi-trip operation and staring imaging operations, the spacecraft body is commonly subject to high-rate motion. This posts challenges for a spacecraft attitude determination subsystem called Gyro Stellar Inertial Attitude Estimate (GS IAE), which employs gyros and star sensors to maintain the required attitude knowledge, since star trackers will severely degrade attitude estimation accuracies when the spacecraft is subject to high-rate motion. This paper analyses the star motion-induced errors for a typical star tracker, models the star motion-induced errors to assess the performance impact on the attitude estimation accuracy, and investigates the adaptive extended Kalman filter design in the GS IAE while evaluating its effectiveness.

PSATdb: a database of mitochondrial common, polymorphic, and unique microsatellites.

Microsatellites, also termed as simple sequence repeats, are repetitive tracts in a DNA sequence, typically consisting of one to six nucleotides. These repeats are found in all genomes and play key roles in phylogeny and species identification. Microsatellites are highly polymorphic, and their length may differ from species to species. There are several online resources dedicated to mitochondria; however, comprehensive information is not available about the length variation of mitochondrial microsatellites. Therefore, to explore it between species among a genus, we have developed a database named pSATdb (polymorphic microSATellites database; https://lms.snu.edu.in/pSATdb/). pSATdb contains 28,710 perfect microsatellites identified across 5,976 mitochondrial genome (mt-genome) sequences from 1,576 genera which includes 1,535 (5,846 mt-genome) and 41 (130 mt-genome) genera of Metazoa and Viridiplantae, respectively. pSATdb is the only database which provides genus-wise information about the length variation of mitochondrial microsatellites. Because of the emerging role of microsatellites in genomics studies, the identified common, polymorphic, and unique microsatellites stored in pSATdb will be effectively useful in various studies including genetic diversity, mapping, marker-assisted selection, and comparative population studies.

MutL homolog 1 methylation and microsatellite instability in sporadic colorectal tumors among Filipinos.

BACKGROUNDColorectal cancer (CRC) ranks third in terms of incidence and second in mortality worldwide. In CRC, the silencing of mismatch repair genes, including the mutL homolog 1 (hMLH1) has been linked to microsatellite instability (MSI), the lengthening or shortening of microsatellite repeats. Very limited data have been presented so far on the link of hMLH1 methylation and MSI in Southeast Asia populations with sporadic CRC, and on its clinical significance.AIMTo investigate the significance of the MSI status and hMLH1 methylation in CRC Filipino patients.METHODSFifty-four sporadic CRC patients with complete clinical data were included in this study. Genomic DNA from CRC tumor biopsies and their normal tissue counterparts were profiled for MSI by high resolution melting (HRM) analysis using the Bethesda Panel of Markers (BAT25, BAT26, D2S123, D5S346, and D17S250). hMLH1 methylation screening was performed using bisulfite conversion and methylation specific polymerase chain reaction. Statistical analysis was conducted to calculate their associations to clinicopathological characteristics and survival relevance (Kaplan-Meier curves and the log-rank test).RESULTS hMLH1 methylation was observed in 9% and 35% of CRC and normal samples, respectively. Higher incidence of consistently methylated hMLH1 found in both normal and CRC was noticed for relation to location of tumor (P < 0.05). As for MSI status, D2S123 the most common unstable microsatellite and MSI-high (MSI-H) was the most common MSI profile, counted for 46% and 50% of normal and CRC tissues, respectively. The presence of MSI-low (MSI-L) and microsatellite stable (MSS) was 43% and 11% for normal, and 31% and 19% for CRC samples. The mean month of patients’ survival was shorter in patients whose normal and tumor tissues had methylated compared to those with unmethylated hMLH1 and with MSI-H compared to those with MSI-L/MSS (P < 0.05). This was supported by significant difference in Kaplan-Meier with log-rank analysis. This data indicated that hMLH1 methylation and high MSI status have prognostic value.CONCLUSIONThis study showed the clinical significance of hMLH1 methylation and MSI status in sporadic CRC Filipino patients, especially in the normal part of the tumor.

Russian-olive (Elaeagnus angustifolia) genetic diversity in the western United States and implications for biological control

AbstractInvasions can be genetically diverse, and that diversity may have implications for invasion management in terms of resistance or tolerance to control methods. We analyzed the population genetics of Russian-olive (Elaeagnus angustifoliaL.), an ecologically important and common invasive tree found in many western U.S. riparian areas. We found three cpDNA haplotypes and, using 11 microsatellite loci, identified three genetic clusters in the 460 plants from 46 populations in the western United States. We found high levels of polymorphism in the microsatellites (5 to 15 alleles per locus; 106 alleles total). Our native-range sampling was limited, and we did not find a genetic match for the most common cpDNA invasive haplotype or a strong confirmation of origin for the most common microsatellite genetic cluster. We did not find geographic population structure (isolation by distance) across the U.S. invasion, but we did identify invasive populations that had the most diversity, and we suggest these as choices for initial biological control–release monitoring. Accessions from each genetic cluster, which coarsely represent the range of genetic diversity found in the invasion, are now included in potential classical biological control agent efficacy testing.

Abstract B011: Immunoediting in untreated mismatch repair deficient colorectal cancer

Abstract A subset of colorectal carcinomas (CRC) with high microsatellite instability (MSI-high) have been shown to respond to immune checkpoint blockade, while the much more common microsatellite stable (MSS) CRC do not usually respond to these immunotherapies. Most MSI-high tumors grow progressively despite a high mutational load, suggesting that the cancer may have developed means to escape from immune recognition, which is termed immunoediting. This immunoediting process includes three phases: an initial phase of nascent cancer cell elimination due to high immunogenicity, then a period of equilibrium with the immune response, finally leading to escape of the cancer cells that have developed genetic or epigenetic mechanisms allowing them to grow progressively in an immunocompetent host. It is likely that MSI-high cancers would go through similar processes to eventually escape from immune recognition. To better understand the differential immunoediting in MSI-high and MSS CRC, we combined tumor exome data from 592 cases from The Cancer Genome Atlas (TCGA) with previously published tumor exome data from 619 cases from the Nurses’ Health Study (NHS) and the Health Professionals Follow-up Study to yield an unprecedented sample of 1,211 CRC molecularly characterized cases. The large number of samples, including a very large number of MSI-high cases (N = 179), gives us the power to identify the prevalence of the mechanisms of resistance discussed and thereby prioritize downstream efforts. We utilized the MutSigCV algorithm to identify significantly mutated genes in MSS and MSI-high tumors in the TCGA cohort (496 MSS and 75 MSI-high cases), and the NHS/HPFS cohorts (438 MSS and 104 MSI-high cases). We identified a total of 62 significantly mutated genes, of which 9 were identified in MSS tumors only, 40 in MSI-high tumors only, and 13 in both. Twenty-seven of the 53 significantly mutated genes in MSI-high were novel, likely resulting from a large increase in the number of MSI-high tumors relative to previous studies. We identified 13 significantly mutated genes with a documented role in the immune system, specifically in MSI-high tumors, a subtype shown previously to have a high level of T-cell infiltration; ZFP36L2 and B2M were significantly mutated in both MSS and MSI-high. B2M, HLA-A, and HLA-B are significantly mutated antigen presenting genes that were previously reported to be recurrently mutated in MSI-high. Eleven of the significantly mutated immune-related genes had roles in modulating diverse hematopoietic cell types and effects beyond antigen presentation; those genes include XYLT2, a dendritic cell trafficking gene; ZBTB20, a Toll-like receptor mediating immune response; RNF128, a regulator of IL2 and IL4 mediated T-cell maturation; KLF3, a gene involved in B-cell development; ZFP36L2, a gene involved in thymocyte development; CASP8, a gene involved in innate immunity; and CD58, a gene involved in the activation of both T lymphocytes and NK cells. Together, these mutations indicate positive selection for immune escape through other mechanisms beyond mutations in antigen-presenting machinery. Citation Format: Catherine Grasso, Marios Giannakis, Daniel Wells, David Wheeler, Eve Shinbrot, Syed Zaidi, Jeroen Huyghe, Milan Geybels, Stephen Salipante, Gabriel Abril-Rodriguez, Helena Escuin-Ordinas, Cristina Puig-Saus, Daniel Sanghoon Shin, Shuji Ogino, Antoni Ribas, Ulrike Peters. Immunoediting in untreated mismatch repair deficient colorectal cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr B011.

Molecular Epidemiology and Antifungal Susceptibility of Candida glabrata in China (August 2009 to July 2014): A Multi-Center Study.

Candida glabrata is an increasingly important cause of invasive candidiasis. In China, relatively little is known of the molecular epidemiology of C. glabrata and of its antifungal susceptibility patterns. Here we studied 411 non-duplicate C. glabrata isolates from 411 patients at 11 hospitals participating in the National China Hospital Invasive Fungal Surveillance Net program (CHIF-NET; 2010-2014). Genotyping was performed using multilocus sequence typing (MLST) employing six genetic loci and by microsatellite analysis. Antifungal susceptibility testing was performed using Sensititre YeastOne™ YO10 methodology. Of 411 isolates, 35 sequence types (ST) were identified by MLST and 79 different genotypes by microsatellite typing; the latter had higher discriminatory power than MLST in the molecular typing of C. glabrata. Using MLST, ST7 and ST3 were the most common STs (66.4 and 9.5% of all isolates, respectively) with 24 novel STs identified; the most common microsatellite types were T25 (30.4% of all isolates) and T31 (12.4%). Resistance to fluconazole (MIC > 32 μg/mL) was seen in 16.5% (68/411) of isolates whilst MICs of >0.5 μg/mL for voriconazole, >2 μg/mL for itraconazole and >2 μg/mL for posaconazole were seen for 28.7, 6.8, and 7.3% of isolates, respectively; 14.8% of all isolates cross-resistant/non-wide-type to fluconazole and voriconazole. Fluconazole resistant rates increased 3-fold over the 5-year period whilst that of isolates with non-WT MICs to voriconazole, 7-fold. All echinocandins exhibited >99% susceptibility rates against all isolates but notably one isolate exhibited multi-drug resistance to the azoles and echinocandins. The study has provided a global picture of the molecular epidemiology and drug resistance rates of C. glabrata in China during the period of the study.

A Clonal Lineage of Fusarium oxysporum Circulates in the Tap Water of Different French Hospitals.

We demonstrated that a clonal lineage of Fusarium oxysporum inhabits the water distribution systems of several French hospitals. This clonal lineage, which appears to be particularly adapted to water networks, represents a potential risk for human infection and raises questions about its worldwide distribution.

Identification of common, unique and polymorphic microsatellites among 73 cyanobacterial genomes.

Microsatellites also known as Simple Sequence Repeats are short tandem repeats of 1-6 nucleotides. These repeats are found in coding as well as non-coding regions of both prokaryotic and eukaryotic genomes and play a significant role in the study of gene regulation, genetic mapping, DNA fingerprinting and evolutionary studies. The availability of 73 complete genome sequences of cyanobacteria enabled us to mine and statistically analyze microsatellites in these genomes. The cyanobacterial microsatellites identified through bioinformatics analysis were stored in a user-friendly database named CyanoSat, which is an efficient data representation and query system designed using ASP.net. The information in CyanoSat comprises of perfect, imperfect and compound microsatellites found in coding, non-coding and coding-non-coding regions. Moreover, it contains PCR primers with 200 nucleotides long flanking region. The mined cyanobacterial microsatellites can be freely accessed at www.compubio.in/CyanoSat/home.aspx. In addition to this 82 polymorphic, 13,866 unique and 2390 common microsatellites were also detected. These microsatellites will be useful in strain identification and genetic diversity studies of cyanobacteria.

Species or Genotypes? Reassessment of Four Recently Described Species of the Ceratocystis Wilt Pathogen, Ceratocystis fimbriata, on Mangifera indica.

Ceratocystis wilt is among the most important diseases on mango (Mangifera indica) in Brazil, Oman, and Pakistan. The causal agent was originally identified in Brazil as Ceratocystis fimbriata, which is considered by some as a complex of many cryptic species, and four new species on mango trees were distinguished from C. fimbriata based on variation in internal transcribed spacer sequences. In the present study, phylogenetic analyses using DNA sequences of mating type genes, TEF-1α, and β-tubulin failed to identify lineages corresponding to the four new species names. Further, mating experiments found that the mango isolates representing the new species were interfertile with each other and a tester strain from sweet potato (Ipomoea batatas), on which the name C. fimbriata is based, and there was little morphological variation among the mango isolates. Microsatellite markers found substantial differentiation among mango isolates at the regional and population levels, but certain microsatellite genotypes were commonly found in multiple populations, suggesting that these genotypes had been disseminated in infected nursery stock. The most common microsatellite genotypes corresponded to the four recently named species (C. manginecans, C. acaciivora, C. mangicola, and C. mangivora), which are considered synonyms of C. fimbriata. This study points to the potential problems of naming new species based on introduced genotypes of a pathogen, the value of an understanding of natural variation within and among populations, and the importance of phenotype in delimiting species.

Patterns of domestication in the Ethiopian oil-seed crop noug (Guizotia abyssinica).

Noug (Guizotia abyssinica) is a semidomesticated oil-seed crop, which is primarily cultivated in Ethiopia. Unlike its closest crop relative, sunflower, noug has small seeds, small flowering heads, many branches, many flowering heads, and indeterminate flowering, and it shatters in the field. Here, we conducted common garden studies and microsatellite analyses of genetic variation to test whether high levels of crop–wild gene flow and/or unfavorable phenotypic correlations have hindered noug domestication. With the exception of one population, analyses of microsatellite variation failed to detect substantial recent admixture between noug and its wild progenitor. Likewise, only very weak correlations were found between seed mass and the number or size of flowering heads. Thus, noug's ‘atypical’ domestication syndrome does not seem to be a consequence of recent introgression or unfavorable phenotypic correlations. Nonetheless, our data do reveal evidence of local adaptation of noug cultivars to different precipitation regimes, as well as high levels of phenotypic plasticity, which may permit reasonable yields under diverse environmental conditions. Why noug has not been fully domesticated remains a mystery, but perhaps early farmers selected for resilience to episodic drought or untended environments rather than larger seeds. Domestication may also have been slowed by noug's outcrossing mating system.

Genetic characterization and relationships of traditional grape cultivars from Serbia

Reference genetic profiles were generated for 12 traditional grapevine cultivars of Serbia through a genotyping approach that included the core set of 9 SSR markers for genetic identification and further 13 common microsatellites for strengthening genetic relationship analysis. Consistent matching with SSR markers of grapevines cultivated in neighbouring countries or maintained in European germplasm collections was found for most of the genotypes, suggesting possible synonyms and revealing that 'Muskat Krokan' corresponds to 'Muscat fleur d’Oranger' and two 'Tamjanika' cultivars are identical to 'Moscato Giallo' and 'Moscato Rosa'. When compared with germplasm representing the classical eco-geographic grouping of grapevine cultivars, Serbian non-Muscat genotypes clustered within the Convar pontica subconvar balcanica taxon thus supporting their indigenous origin.

An exotic herbivorous insect drives the evolution of resistance in the exotic perennial herb Solidago altissima

Invasive plants often experience rapid changes in biological interactions by escaping from their original herbivores at their new habitats, and sometimes re‐associating with those herbivores afterwards. However, little is known about whether the temporal changes in herbivorous impact work as a selective agent for defensive traits of invaded plants. Solidago altissima (goldenrod) is a North American perennial that has widely invaded abandoned fields in Japan. Recently, an herbivorous insect Corythucha marmorata (lace bug), an exotic insect also from North America, which was first recorded in 2000 in Japan, has been expanding its habitat on S. altissima populations in Japan. In this study, we investigated whether the invasion of C. marmorata had a selective impact on the defensive traits of S. altissima, by conducting a field survey, a common garden experiment and microsatellite analysis. We compared quantitative genetic differentiation of traits (resistance, growth, and reproduction) and neutral molecular differentiation among 16 S. altissima populations with different establishment years of C. marmorata.The common garden experiment, in which plants were grown in a greenhouse and treated to either C. marmorata herbivory or no herbivory, revealed the presence of higher resistance, sexual reproduction, and asexual (rhizome) reproduction in populations subjected to a longer history of C. marmorata pressure. Such phenotypic variability among establishment years of lace bugs was likely driven by natural selection rather than stochastic events such as genetic drift and founder effects. In addition, when plants were exposed to lace bug herbivory, resistance had a positive relationship with sexual and asexual reproduction, although no relationship was found when plants were free from herbivory. These findings suggest that defensive traits in S. altissima have evolved locally in the last decade in response to the selective pressure of C. marmorata.

Genetic structure of the critically endangered Red‐headed Wood Pigeon Columba janthina nitens and its implications for the management of threatened island populations

The Red‐headed Wood Pigeon Columba janthina nitens is endemic to the Ogasawara Islands, an oceanic island chain located 1000 km south of the main islands of Japan. The subspecies is at high risk of extinction because of its small population size and restricted habitat range. We undertook genetic analyses of this pigeon using sequences of a portion of the mitochondrial control region and five microsatellite markers to estimate the genetic characteristics of two wild populations from the Bonin and Volcano Islands, as well as one captive breeding population. The genetic diversity of the wild individuals was exceptionally low in both the mitochondria (nucleotide diversity = 0.00105) and at the microsatellite (3.2 alleles per locus and HE = 0.12) loci. Higher numbers of microsatellite genotypes were observed in the Volcano Islands population than in the Bonin Islands population, which may be because of the relatively low impact of human disturbance. The most common mitochondrial haplotypes and microsatellite alleles observed in the two wild populations were completely fixed in the captive population. Our results suggest that the genetic diversity of the captive population needs to be increased. However, introduction of a wild individual into a captive population can lead to a decreased genetic diversity in the wild population and therefore should be done with caution. The genetic differentiation between the Bonin and the Volcano island groups was low, and the populations of the two island groups should be regarded as a single evolutionarily significant unit. However, special consideration is required for habitat conservation in the Volcano Islands, which may be functioning as a sanctuary for the Red‐headed Wood Pigeon. For the long‐term conservation of threatened bird species that live on remote oceanic islands, determination of management units considering gene flow caused by their flying capacity and maintenance of genetically suitable wild and captive populations are essential.

Bax 2 Family Alternative Splicing Salvages Bax Microsatellite-Frameshift Mutations

Mutation or aberrant splicing can interrupt gene expression. Tumor suppressor Bax is one of the susceptible genes prone to microsatellite frameshifting mutations in coding regions. As a result, tumors exhibiting microsatellite instability (MSI) often present a "Bax-negative" phenotype. We previously reported that some Bax-negative cells in fact contain a functional Bax isoform (BaxΔ2), generated when unique alternative splicing "salvages" the shifted reading frame introduced by a microsatellite mutation. Here we compared Bax alternative splicing profiles in a range of cell lines and primary tumors with and without Bax microsatellite mutations. We found that MSI tumors exhibit a high Bax alternative splicing frequency, especially in exon 2, and produce a family of alternatively spliced isoforms that retain many important Bax functional domains. Surprisingly, these BaxΔ2 family isoforms can rescue Bax from all common microsatellite frameshift mutations. Production of BaxΔ2 requires specific cis mutations, while trans components are not cell-type specific. Furthermore, all BaxΔ2 family isoforms are more potent cell death inducers than the parental Bax without directly targeting mitochondria. These results indicate that the BaxΔ2 family can potentially salvage Bax tumor suppressor expression otherwise lost to mutation.

Genetic diversity and population structure assessed by SSR and SNP markers in a large germplasm collection of grape

BackgroundThe economic importance of grapevine has driven significant efforts in genomics to accelerate the exploitation of Vitis resources for development of new cultivars. However, although a large number of clonally propagated accessions are maintained in grape germplasm collections worldwide, their use for crop improvement is limited by the scarcity of information on genetic diversity, population structure and proper phenotypic assessment. The identification of representative and manageable subset of accessions would facilitate access to the diversity available in large collections. A genome-wide germplasm characterization using molecular markers can offer reliable tools for adjusting the quality and representativeness of such core samples.ResultsWe investigated patterns of molecular diversity at 22 common microsatellite loci and 384 single nucleotide polymorphisms (SNPs) in 2273 accessions of domesticated grapevine V. vinifera ssp. sativa, its wild relative V. vinifera ssp. sylvestris, interspecific hybrid cultivars and rootstocks. Despite the large number of putative duplicates and extensive clonal relationships among the accessions, we observed high level of genetic variation. In the total germplasm collection the average genetic diversity, as quantified by the expected heterozygosity, was higher for SSR loci (0.81) than for SNPs (0.34). The analysis of the genetic structure in the grape germplasm collection revealed several levels of stratification. The primary division was between accessions of V. vinifera and non-vinifera, followed by the distinction between wild and domesticated grapevine. Intra-specific subgroups were detected within cultivated grapevine representing different eco-geographic groups. The comparison of a phenological core collection and genetic core collections showed that the latter retained more genetic diversity, while maintaining a similar phenotypic variability.ConclusionsThe comprehensive molecular characterization of our grape germplasm collection contributes to the knowledge about levels and distribution of genetic diversity in the existing resources of Vitis and provides insights into genetic subdivision within the European germplasm. Genotypic and phenotypic information compared in this study may efficiently guide further exploration of this diversity for facilitating its practical use.

Population structure of Sclerotinia sclerotiorum in crop and wild hosts in the UK

Sclerotinia sclerotiorum is an important pathogen of many crop plants which also infects wild hosts. The population structure of this fungus was studied for different crop plants and Ranunculus acris (meadow buttercup) in the UK using eight microsatellite markers and sequenced sections of the intergenic spacer (IGS) region of the rRNA gene and the elongation factor 1‐alpha (EF) gene. A total of 228 microsatellite haplotypes were identified within 384 isolates from 12 S. sclerotiorum populations sampled in England and Wales. One microsatellite haplotype was generally found at high frequency in each population and was distributed widely across different hosts, locations and years. Fourteen IGS and five EF haplotypes were found in the 12 populations, with six IGS haplotypes and one EF haplotype exclusive to buttercup. Analysis of published sequences for S. sclerotiorum populations from the USA, Canada, New Zealand and Norway showed that three of the IGS haplotypes and one EF haplotype were widely distributed, while eight IGS haplotypes were only found in the UK. Although common microsatellite and IGS/EF haplotypes were found on different hosts in the UK, there was evidence of differentiation, particularly for one isolated population on buttercup. However, overall there was no consistent differentiation of S. sclerotiorum populations from buttercup and crop hosts. Sclerotinia sclerotiorum therefore has a multiclonal population structure in the UK and the wide distribution of one microsatellite haplotype suggests spatial mixing at a national scale. The related species S. subarctica was also identified in one buttercup population.

New highly polymorphic microsatellite loci for the Galápagos marine iguana, Amblyrhynchus cristatus

We describe the development and characterisation of six new dinucleotide motif microsatellite loci for populations of marine iguanas (Amblyrhynchus cristatus), endemic to the Galápagos archipelago. Primers were based on microsatellite-bearing sequences and initially developed using universally labelled primers. When analysed across 5 populations (representing 150 individuals), new loci displayed, on average, high levels of genetic diversity (range: 2-13 alleles, mean: 5.73) and values of heterozygosity (range: 0.0-0.906, mean: 0.605). No consistent deviations from Hardy-Weinberg equilibrium or significant linkage disequilibrium were observed, and all loci were shown to be free of common microsatellite errors. Utilising the 13 previously available microsatellite loci for this species, we describe here four multiplex combinations for the successful amplification of 19 microsatellite loci for marine iguanas. This powerful set of highly polymorphic markers will allow researchers to explore future questions regarding the ecology, evolution, and conservation of this unique species.