Allele Length Research Articles

C9orf72 hexanucleotide repeat length in older population: normal variation and effects on cognition

The hexanucleotide repeat expansion in C9orf72 is a common cause of amyotrophic lateral sclerosis/frontotemporal dementia and also rarely found in other psychiatric and neurodegenerative conditions. Alleles with >30 repeats are often considered an expansion, but the pathogenic repeat length threshold is still unclear. It is also unclear whether intermediate repeat length alleles (often defined either as 7–30 or 20–30 repeats) have clinically significant effects. We determined the C9orf72 repeat length distribution in 3142 older Finns (aged 60–104 years). The longest nonexpanded allele was 45 repeats. We found 7–45 repeats in 1036/3142 (33%) individuals, 20–45 repeats in 56/3142 (1.8%), 30–45 repeats in 12/3142 (0.38%), and expansion (>45 repeats) in 6/3142 (0.19%). There was no apparent clustering of neurodegenerative or psychiatric diseases in individuals with 30–45 repeats indicating that 30–45 repeats are not pathogenic. None of the 6 expansion carriers had a diagnosis of amyotrophic lateral sclerosis/frontotemporal dementia but 4 had a diagnosis of a neurodegenerative or psychiatric disease. Intermediate length alleles (categorized as 7–45 and 20–45 repeats) did not associate with Alzheimer's disease or cognitive impairment.

A multi-task convolutional deep neural network for variant calling in single molecule sequencing

The accurate identification of DNA sequence variants is an important, but challenging task in genomics. It is particularly difficult for single molecule sequencing, which has a per-nucleotide error rate of ~5–15%. Meeting this demand, we developed Clairvoyante, a multi-task five-layer convolutional neural network model for predicting variant type (SNP or indel), zygosity, alternative allele and indel length from aligned reads. For the well-characterized NA12878 human sample, Clairvoyante achieves 99.67, 95.78, 90.53% F1-score on 1KP common variants, and 98.65, 92.57, 87.26% F1-score for whole-genome analysis, using Illumina, PacBio, and Oxford Nanopore data, respectively. Training on a second human sample shows Clairvoyante is sample agnostic and finds variants in less than 2 h on a standard server. Furthermore, we present 3,135 variants that are missed using Illumina but supported independently by both PacBio and Oxford Nanopore reads. Clairvoyante is available open-source (https://github.com/aquaskyline/Clairvoyante), with modules to train, utilize and visualize the model.

The fragile X CGG repeat shows a marked level of instability in hereditary non-polyposis colorectal cancer patients.

The allelic variation of the FMR1 CGG repeat was investigated by small-pool PCR in nonneoplastic peripheral blood leukocytes from HNPCC patients and matched controls for similar CGG repeat lengths. The allelic variation for repeat lengths appears to be roughly twice as frequent in HNPCC patients as in controls, especially when patients are mutated in hMLH1. There are more expansions in HNPCC patients (42%) than in controls (20%) but this difference is statistically borderline. The mean length of expansions relative to the genuine size did not differ in HNPCC patients or controls (respectively 17% and 20% of the constitutional allelic length). The reported data suggest that instability within nonneoplastic cells of a subset of HNPCC patients might be one mechanism for transition from normal to the premutation range of the FMR1 CGG repeat.

Trinucleotide repeat expansion length as a predictor of the clinical progression of Fuchs' Endothelial Corneal Dystrophy.

PurposeTo determine if CTG18.1 TNR expansion length prognosticates the clinical progression of Fuchs’ Endothelial Corneal Dystrophy (FECD).MethodsThis was a prospective cohort study. A total of 51 patients with newly diagnosed FECD were recruited and followed-up over a period of 12 years, from November 2004 to April 2016. Baseline clinical measurements included central corneal thickness (CCT), endothelial cell density (ECD) and CTG18.1 TNR expansion length from peripheral leukocytes, with yearly repeat measurements of CCT and ECD. A patient was defined to have experienced significant clinical progression and to have developed Threshold Disease if any of these criteria were fulfilled in either eye: a) CCT increased to >700μm, b) ECD decreased to <700 cells/mm2, or c) underwent keratoplasty for treatment of FECD.ResultsPatients were categorized as having at least one allele whose maximum allele length was equal to or greater than 40 repeats (L≥40, n = 22, 43.1%), or having both alleles shorter than 40 repeats (L<40). Threshold Disease rates at the 5-year time point were 87.5% for the L≥40 group and 47.8% for the L<40 group (p = 0.012). This difference narrowed and was no longer statistically significant at the 8-years (92.9% vs 78.9%, p = 0.278) and 10-years (92.9% vs 84.2%, p = 0.426) time points.ConclusionsL≥40 patients are at greater risk of FECD progression and development of Threshold Disease within the first 5 years following diagnosis.

\xb5LAS: Sizing of expanded trinucleotide repeats with femtomolar sensitivity in less than 5\u2009minutes

We present µLAS, a lab-on-chip system that concentrates, separates, and detects DNA fragments in a single module. µLAS speeds up DNA size analysis in minutes using femtomolar amounts of amplified DNA. Here we tested the relevance of µLAS for sizing expanded trinucleotide repeats, which cause over 20 different neurological and neuromuscular disorders. Because the length of trinucleotide repeats correlates with the severity of the diseases, it is crucial to be able to size repeat tract length accurately and efficiently. Expanded trinucleotide repeats are however genetically unstable and difficult to amplify. Thus, the amount of amplified material to work with is often limited, making its analysis labor-intensive. We report the detection of heterogeneous allele lengths in 8 samples from myotonic dystrophy type 1 and Huntington disease patients with up to 750 CAG/CTG repeats in five minutes or less. The high sensitivity of the method allowed us to minimize the number of amplification cycles and thus reduce amplification artefacts without compromising the detection of the expanded allele. These results suggest that µLAS can speed up routine molecular biology applications of repetitive sequences and may improve the molecular diagnostic of expanded repeat disorders.

Correlation between Serotonin Transporter Promoter Gene Polymorphism and PTSD in Children

Objective: To study the association between post-traumatic stress disorder (PTSD) and serotonin transporter promoter (5-HTTLPR) gene polymorphism in Han children in Hainan; to explore the genetic mechanism of PTSD in children. Methods: 50 patients with post-traumatic stress disorder and healthy children in Han nationality in Hainan were selected. Detection of 5-HTTLPR gene polymorphism by polymerase chain reaction (PCR) and amplified fragment length polymorphism, the genotype and allele frequencies were analyzed using a case-control association analysis method. Results: There were 4, 14 and 32 cases of LL, SL and SS in the post-traumatic stress disorder group of Hainan Han children, and 13, 20 and 17 cases in the control group. From the perspective of gene frequency, the L gene of post-traumatic stress disorder appeared 22.0%, and S appeared 78.0%. In the control group, L appeared 46.0%, and S appeared 54.0%. There were significant differences in genotype and gene frequency (P < 0.01). Conclusion: The 5-HTTLPR gene polymorphism in Hainan Han children may be associated with post-traumatic stress disorder.

CAG polymorphism in the androgen receptor gene in women may be associated with nodulocystic acne.

IntroductionAcne vulgaris (AV) is a multifactorial, inflammatory disease of the pilosebaceous unit. Hormones play a major role in the pathogenesis of acne. In cases of hyperandrogenism; hirsutism, acne, seborrhoea and alopecia appear in women. However, severe acne can also be seen without evidence of hyperandrogenism. In this case, hypersensitivity of the androgen receptor gene (ARG) encoded in the X chromosome, which is the only receptor for androgens, can be considered. ARG contains a polymorphic CAG triple loop encoding the polyglutamine pathway at the 5’end of exon 1.AimTo investigate CAG repeat polymorphism in the ARG in nodulocystic acne patients in Turkish population.Material and methodsThis prospective clinical study was conducted between 2016 and 2017 in accordance with the tenets of the Declaration of Helsinki. DNA isolation from blood was performed using the RTA® Genomic DNA Isolation Kit. The fragment lengths obtained from the device to determine CAG repeat numbers were analysed based on –288 bp length 22 CAG repeat content.ResultsA total of 199 subjects; 100 patients (51 males, 49 females) and 99 controls (49 males, 50 females) were included in the study. The mean allele length in the patient group was 19.34; and 19.7 in the control group. There was a statistically significant difference between female patients and the control group, when the patients and control groups were compared by gender (p = 0.0059).ConclusionsThe CAG trinucleotide repeat count in the ARG may be associated with acne, without hirsutism findings.

The association between repeat number in C9orf72 and phenotypic variability in Turkish patients with frontotemporal lobar degeneration

Frontotemporal lobar degeneration (FTLD) describes a group of progressive brain disorders. The expansion of a noncoding GGGGCC (G4C2) hexanucleotide repeat in the C9orf72 gene is a major cause of both familial FTLD and amyotrophic lateral sclerosis. The aim of this study was to determine the prevalence of C9orf72 G4C2-repeat expansion in a Turkish population with FTLD and to determine its effects on the phenotype. The G4C2 expansion in the C9orf72 gene was analyzed in 100 cases of FTLD without mutations of the MAPT, PGRN, CHMP2B, VCP, TARDBP, and FUS genes and 100 age-matched healthy controls by using repeat-primed polymerase chain reaction and fragment length analysis techniques. A possible pathogenic repeat (≥30) was found in one of the familial cases (1/33), but none of the sporadic cases. The difference in the allele length between the cases and controls was statistically significant (p < 0.01). Intermediate (20–30) repeats were detected in 4% of our cases. Patients with psychotic symptoms appear to be enriched for intermediate and possibly pathogenic repeats. To determine whether the intermediate and ≥30-repeat allele carriers shared the C9orf72 risk haplotype, we examined rs4879515 and rs3849942 in all samples and family members of patients with possibly pathogenic alleles. We identified at least one risk allele for each single-nucleotide polymorphism in all intermediate and possibly pathogenic repeat carriers. We observed that ≥8 unit repeats were strongly correlated with the tagging risk alleles for both single-nucleotide polymorphisms (p < 0.001). To our knowledge, this is the first study to evaluate C9orf72 G4C2 repeats in Turkish patients with FTLD. The present findings suggest that pathogenic expansions of the C9orf72 repeat are uncommon in Turkish patients with FTLD, but intermediate repeats may be a risk factor for FTLD and act as a genetic modifying factor for psychotic symptoms.

Analysis of microsatellite loci in tree of heaven (Ailanthus altissima (Mill.) Swingle) using SSR-GBS

Microsatellite markers are still the marker of choice for many research questions in the field of forest genetics. However, the number of available markers is often low for species that have not been studied intensively like the tree of heaven (Ailanthus altissima). During the last decade, next-generation sequencing (NGS) has offered advanced techniques for efficiently identifying microsatellite markers and accurately genotyping samples. Here, we identify new microsatellite markers for the tree of heaven by applying an NGS-based method using the Illumina MiSeq platform. NGS technology was proved to be an effective method for fast and cost-efficient identification of microsatellite markers by implementing a genotyping-by-sequencing approach based on Illumina amplicon sequencing (SSR-GBS). We screened three populations from Eastern Austria for genetic variation at 19 newly identified microsatellite loci. We tested two different genotyping approaches: (1) considering only allele lengths (forming a so-called “allele length dataset”), (2) taking also single nucleotide polymorphisms (SNPs) within the amplified fragments into account (forming a so-called “SNP dataset”). The results revealed higher values for all genetic diversity parameters, as well as a better resolution of genetic assignment, when the latter approach was followed. Thus, by taking advantage of sequence information which is provided by SSR-GBS, one may achieve considerable gains in performance using the same marker set. The developed markers provide a cost-efficient tool for genotyping populations of tree of heaven and the approach presented here promises to be of high value for medium throughput genotyping applications in non-model forest tree species. We will use this method to widen the perspectives for further population genetic investigations of the tree of heaven.

A patient-derived cellular model for Huntington's disease reveals phenotypes at clinically relevant CAG lengths.

The huntingtin protein participates in several cellular processes that are disrupted when the polyglutamine tract is expanded beyond a threshold of 37 CAG DNA repeats in Huntington’s disease (HD). Cellular biology approaches to understand these functional disruptions in HD have primarily focused on cell lines with synthetically long CAG length alleles that clinically represent outliers in this disease and a more severe form of HD that lacks age onset. Patient-derived fibroblasts are limited to a finite number of passages before succumbing to cellular senescence. We used human telomerase reverse transcriptase (hTERT) to immortalize fibroblasts taken from individuals of varying age, sex, disease onset, and CAG repeat length, which we have termed TruHD cells. TruHD cells display classic HD phenotypes of altered morphology, size and growth rate, increased sensitivity to oxidative stress, aberrant adenosine diphosphate/adenosine triphosphate (ADP/ATP) ratios, and hypophosphorylated huntingtin protein. We additionally observed dysregulated reactive oxygen species (ROS)-dependent huntingtin localization to nuclear speckles in HD cells. We report the generation and characterization of a human, clinically relevant cellular model for investigating disease mechanisms in HD at the single-cell level, which, unlike transformed cell lines, maintains functions critical for huntingtin transcriptional regulation and genomic integrity.

Potential highly polymorphic short tandem repeat markers for enhanced forensic identity testing

Due to their polymorphic nature, short tandem repeats (STRs) are well-studied and routinely used genetic markers for forensic DNA typing. However, even the largest STR multiplexes are limited in their ability to parse out individuals in a DNA mixture sample, due to alleles shared by size detected by capillary electrophoresis and challenges in resolving minor alleles from stutter, and inherent heterozygote imbalance. In this study, STRs were explored in public datasets that displayed sequence variation and may have limited allele length spread. STRs were first selected using fundamental criteria of high heterozygosity, tetra-, penta-, or hexanucleotide repeat length, and overall relative narrow allele spread (based on length). All candidates were further scrutinized for chemistry compatibility. The resulting STRs were multiplexed and sequenced by massively parallel sequencing in a limited sample population set. Each candidate STR was evaluated for analytical performance and desired biological properties. The findings presented describe a refined set of 53 potential highly polymorphic STR markers (high sequence diversity and heterozygosity; reduced allele spread) that may be suitable to supplement the current core marker set(s) for possible enhanced characterization of complex DNA profiles.

Instability of TCF4 Triplet Repeat Expansion With Parent-Child Transmission in Fuchs' Endothelial Corneal Dystrophy.

PurposeFuchs' endothelial corneal dystrophy (FECD) caused by the CTG triplet repeat expansion in the TCF4 gene (CTG18.1 locus) is the most common repeat expansion disorder. Intergenerational instability of expanded repeats and clinical anticipation are hallmarks of other repeat expansion disorders. In this study, we examine stability of triplet repeat allele length and FECD disease severity in parent–child transmission of the expanded CTG18.1 allele.MethodsWe studied 44 parent–child transmissions of the mutant expanded CTG18.1 allele from 26 FECD families. The CTG18.1 polymorphism was genotyped using short tandem repeat analysis, triplet repeat primed PCR assay, and Southern blot analysis. FECD severity was assessed using modified Krachmer grading (KG) system. Triplet repeat length of mutant allele and KG severity were compared between generations.ResultsInstability of the expanded allele was seen in 14 of 44 (31.8%) parent–child transmissions, and the likelihood of an unstable event increased with the size of the parental allele (newcommand{bialpha}{boldsymbol{alpha}}newcommand{bibeta}{boldsymbol{beta}}newcommand{bigamma}{boldsymbol{gamma}}newcommand{bidelta}{boldsymbol{delta}}newcommand{bivarepsilon}{boldsymbol{varepsilon}}newcommand{bizeta}{boldsymbol{zeta}}newcommand{bieta}{boldsymbol{eta}}newcommand{bitheta}{boldsymbol{theta}}newcommand{biiota}{boldsymbol{iota}}newcommand{bikappa}{boldsymbol{kappa}}newcommand{bilambda}{boldsymbol{lambda}}newcommand{bimu}{boldsymbol{mu}}newcommand{binu}{boldsymbol{nu}}newcommand{bixi}{boldsymbol{xi}}newcommand{biomicron}{boldsymbol{micron}}newcommand{bipi}{boldsymbol{pi}}newcommand{birho}{boldsymbol{rho}}newcommand{bisigma}{boldsymbol{sigma}}newcommand{bitau}{boldsymbol{tau}}newcommand{biupsilon}{boldsymbol{upsilon}}newcommand{biphi}{boldsymbol{phi}}newcommand{bichi}{boldsymbol{chi}}newcommand{bipsi}{boldsymbol{psi}}newcommand{biomega}{boldsymbol{omega}}P = 5.9 times {10^{ - 3}}). A tendency for contraction was seen in transmission of large alleles (repeat length > 120), whereas intermediate alleles (repeat length between 77 and 120) had predilection for further expansion (newcommand{bialpha}{boldsymbol{alpha}}newcommand{bibeta}{boldsymbol{beta}}newcommand{bigamma}{boldsymbol{gamma}}newcommand{bidelta}{boldsymbol{delta}}newcommand{bivarepsilon}{boldsymbol{varepsilon}}newcommand{bizeta}{boldsymbol{zeta}}newcommand{bieta}{boldsymbol{eta}}newcommand{bitheta}{boldsymbol{theta}}newcommand{biiota}{boldsymbol{iota}}newcommand{bikappa}{boldsymbol{kappa}}newcommand{bilambda}{boldsymbol{lambda}}newcommand{bimu}{boldsymbol{mu}}newcommand{binu}{boldsymbol{nu}}newcommand{bixi}{boldsymbol{xi}}newcommand{biomicron}{boldsymbol{micron}}newcommand{bipi}{boldsymbol{pi}}newcommand{birho}{boldsymbol{rho}}newcommand{bisigma}{boldsymbol{sigma}}newcommand{bitau}{boldsymbol{tau}}newcommand{biupsilon}{boldsymbol{upsilon}}newcommand{biphi}{boldsymbol{phi}}newcommand{bichi}{boldsymbol{chi}}newcommand{bipsi}{boldsymbol{psi}}newcommand{biomega}{boldsymbol{omega}}P = 1.3 times {10^{ - 3}}). Although we noted increased KG severity in the offspring in three pairs, none of these transmissions were associated with allele instability.ConclusionsWe observed instability of the TCF4 triplet repeat expansion in nearly a third of parent–child transmissions. Large mutant CTG18.1 alleles are prone to contraction, whereas intermediate mutant alleles tend to expand when unstably transmitted. Intergenerational instability of TCF4 repeat expansion has implications on FECD disease inheritance.

INDEL Length and Haplotypes in the β-Synuclein Gene: A Key to Differentiate Dementia with Lewy Bodies?

Lewy body diseases (LBD) include Parkinson's disease (PD) and dementia with Lewy bodies (DLB) and together with Alzheimer's disease (AD) they show an important neuropathological and clinical overlap. The human alpha- and beta-synuclein genes (SNCA and SNCB) are key factors for the development of Lewy body diseases. Here, we aimed to analyze the genotype distribution of potentially functional SNPs in SNCA and SNCB, perform haplotype analysis for SNCB, and to identify functional insertion and deletion (INDEL) variations within the regulatory region of SNCB which might be responsible for the drastically diminished beta-synuclein levels reported for pure DLB. Thus, we genotyped brain samples from AD, DLB, PD, and healthy controls for two SNCA and four SNCB SNPs. We also analyzed INDEL variations upstream of SNCB, determined SNCB expression levels, and correlated INDEL lengths with expression levels. Applying Fisher's exact, chi-square, ANOVA tests, and the ΔΔCt method, we found disease-specific genotype distribution of SNCA and SNCB SNPs. Additionally, we identified three INDEL variations upstream of SNCB and showed that the INDEL allele lengths were associated with SNCB expression levels. INDEL alleles associated with low SNCB expression were accumulated in pure DLB. Finally, one major and four minor DLB specific SNCB haplotypes were identified with Haploview and Arlequin. In summary, our study showed that different SNCA and SNCB genotypes are associated with the development of either PD or DLB, and that the frequencies of genotypes associated with low SNCB expression are elevated in DLB.

Premutations of FMR1 CGG repeats are not related to idiopathic premature ovarian failure in Iranian patients: A case control study

Premature ovarian failure (POF) is a reproductive disease which affects 1 in 100 under 40 years women. FMR1 premutation carriers of CGG repeats are supposed to be at increased risk for POF. We have examined the 5′UTR region of the gene to find any association between the repeat size and the disease etiology in Iranian population. 30 women with early idiopathic POF and 30 fertile control women were selected. We used triplet repeat primed PCR (TP PCR) assay and gene-specific primers to amplify the CpG Island of FMR1 gene promoter region. The amplification results were analyzed by capillary electrophoresis and Gene Marker software. Among 30 patients, two had intermediate repeat size, one had premutation and the rest had CGG repeat of the normal range. Two of controls had intermediate repeats and none had a premutation. Two groups had significant differences in the repeat number average (p = 0.007) and in the average length of the smallest allele (p < 0.001), but had no promising difference in average length of the longest allele (p = 0.453).Although the two groups showed a significant difference in the length of alleles, their length was within normal range. Perhaps the polymorphism, in connection with the genome's entire network, has been involved in the development of the disease, or there has been a fundamentally different mechanism for the disease in Iranian population. A larger number of Iranian POF patients should be investigated for any probable relationship between the CGG triplet repeat length and the etiology of the disease.

Development of a massively parallel sequencing assay for investigating sequence polymorphisms of 15 short tandem repeats in a Chinese Northern Han population.

Massively parallel sequencing (MPS) has been used in forensic genetics in recent years owing to several advantages, e.g. MPS can provide precise descriptions of the repeat allele structure and variation in the repeat-flanking regions, increasing the discriminating power among loci and individuals. However, it cannot be fully utilized unless sufficient population data are available for all loci. Thus, there is a pressing need to perform population studies providing a basis for the introduction of MPS into forensic practice. Here, we constructed a multiplex PCR system with fusion primers for one-directional PCR for MPS of 15 commonly used forensic autosomal STRs and amelogenin. Samples from 554 unrelated Chinese Northern Han individuals were typed using this MPS assay. In total, 313 alleles obtained by MPS for all 15 STRs were observed, and the corresponding allele frequencies ranged between 0.0009 and 0.5162. Of all 15 loci, the number of alleles identified for 12 loci increased compared to capillary electrophoresis approaches, and for the following six loci more than double the number of alleles was found: D2S1338, D5S818, D21S11, D13S317, vWA, and D3S1358. Forensic parameters were calculated based on length and sequence-based alleles. D21S11 showed the highest heterozygosity (0.8791), discrimination power (0.9865), and paternity exclusion probability in trios (0.7529). The cumulative match probability for MPS was approximately 2.3157 × 10-20 .

Sequence-based US population data for the SE33 locus.

A set of 1036 U.S. Population Samples were sequenced using the Illumina ForenSeq DNA Signature Prep Kit. This sample set has been highly characterized using a variety of marker systems for human identification. The FASTQ files obtained from a ForenSeq DNA Signature Prep Kit experiment include several STR loci that are not reported in the associated software. These include SE33, DXS8377, DXS10148, DYS456, and DYS461. The sequence variation within the autosomal STR marker SE33 was evaluated using a customized bioinformatic approach to identify and characterize the locus in the 1036 data set. The analysis identified 53 unique alleles by length and 264 by sequence. An additional 10 alleles were detected when selected extended flanking regions were examined to resolve discordances. Allele frequencies and SE33 sequence motif patterns are reported for the 1036 data set. The comparison of numerical allele calls derived from sequence data to the allele calls obtained from commercial capillary electrophoresis-based STR typing kits resulted in 100% concordance, after manual data review and confirmation sequencing of three flanking region deletions. The analysis of this data set involved significant manual sequence curation and information support from length-based genotypes to ensure high confidence in the sequence-based allele calls. The challenges of interpreting the sequence data for SE33 consisted of high sequence noise, allele-size dependent variance in coverage, and heterozygote imbalance. As allele length increased, sequence depth of coverage and quality decreased at the terminal end. Accordingly, heterozygous genotype imbalance increased in proportion to increased distance between alleles.

RNA-based FLT3-ITD allelic ratio is associated with outcome and ex vivo response to FLT3 inhibitors in pediatric AML

AbstractControversy exists whether internal tandem duplication of FMS-like tyrosine kinase 3 (FLT3-internal tandem duplication [ITD]) allelic ratio (AR) and/or length of the ITD should be taken into account for risk stratification of pediatric acute myeloid leukemia (AML) and whether it should be measured on RNA or DNA. Moreover, the ITD status may be of relevance for selecting patients eligible for FLT3 inhibitors. Here, we included 172 pediatric AML patients, of whom 36 (21%) harbored FLT3-ITD as determined on both RNA and DNA. Although there was a good correlation between both parameters ARspearman = 0.62 (95% confidence interval, 0.22-0.87) and ITDlengthspearman = 0.98 (95% confidence interval, 0.90-1.00), only AR ≥ 0.5 and length ≥48 base pairs (bps) based on RNA measurements were significantly associated with overall survival (AR: Plogrank = .008; ITDlength: Plogrank = .011). In large ITDs (>156 bp on DNA) a remarkable 90-bp difference exists between DNA and RNA, including intron 14, which is spliced out in RNA. Ex vivo exposure (n = 30) to FLT3 inhibitors, in particular to the FLT3-specific inhibitor gilteritinib, showed that colony-forming capacity was significantly more reduced in FLT3-ITD-AR ≥ 0.5 compared with ITD-AR-low and ITD− patient samples (P < .001). RNA-based FLT3-ITD measurements are recommended for risk stratification, and the relevance of AR regarding eligibility for FLT3-targeted therapy warrants further study.

Intermediate-length CAG repeat in ATXN2 is associated with increased risk for amyotrophic lateral sclerosis in Brazilian patients.

Intermediate-length cytosine-adenine-guanine nucleotide repeat expansions in the ATXN2 gene (which encodes for the protein Ataxin-2) have been linked to increased risk for amyotrophic lateral sclerosis (ALS) in different populations. There is no such study in the Brazilian population, which has a mixed ethnic background. We have thus selected 459 patients with ALS (372 Sporadic ALS and 87 Familial ALS) and 468 control subjects from 6 Brazilian centers to investigate this point. We performed polymerase chain reaction to determine the length of the ATXN2 alleles. Polymerase chain reaction products were resolved using capillary electrophoresis on ABI 3500× l capillary sequencer. We found that ATXN2 intermediate-length expansions (larger than 26 repeats) were associated with an increased risk for ALS (odds ratio= 2.56, 95% confidence interval: 1.29-5.08, p= 0.005). Phenotype in patients with and without ATXN2 expansions was similar. Our findings support the hypothesis that ATXN2 plays an important role in the pathogenesis of ALS also in the Brazilian population.

QTL-Seq Identified a Major QTL for Grain Length and Weight in Rice Using Near Isogenic F2 Population

Abstract: Mapping and isolation of quantitative trait loci (QTLs) or genes controlling grain size or weight is very important to uncover the molecular mechanisms of seed development and crop breeding. To identify the QTLs controlling grain size and weight, we developed a near isogenic line F2 (NIL-F2) population, which was derived from a residual heterozygous plant in an F7 generation of recombinant inbred line (RIL). With the completion of more than 30y whole genome re-sequencing of the parents, two DNA bulks for large and small grains, a total of 58.94 Gb clean nucleotide data were generated. A total of 455 262 single nucleotide polymorphisms (SNPs) between the parents were identified to perform bulked QTL-seq. A candidate genomic region containing SNPs strongly associated with grain length and weight was identified from 15 to 20 Mb on chromosome 5. We designated the major QTL in the candidate region as qTGW5.3. Then, qTGW5.3 was further validated with PCR-based conventional QTL mapping method through developing simple sequence repeat and Insertion/Deletion markers in the F2 population. Furthermore, recombinants and the progeny tests delimited the candidate region of qTGW5.3 to 1.13 Mb, flanked by HX5009 (15.15 Mb) and HX5003 (16.28 Mb). A set of NILs, selected from the F2 population, was developed to evaluate the genetic effect of qTGW5.3. Significant QTL effects were detected on grain length, grain width and 1000-grain weight of H12-29 allele with 1.14 mm, -0.11 mm and 3.11 g, which explained 99.64%, 95.51% and 97.32% of the phenotypic variations, respectively.

Targeted exome sequencing reveals homozygous TREM2 R47C mutation presenting with behavioral variant frontotemporal dementia without bone involvement

To identify genes associated with frontotemporal dementia (FTD) in South-East Asia, targeted exome sequencing and C9orf72 genotyping was performed in 198 subjects (52 patients with FTD and 146 healthy controls) who were screened for mutations in 12 FTD-associated genes. We detected a homozygous TREM2 R47C mutation in a patient with behavioral variant FTD without bone cysts or bone-associated phenotype. Two novel nonsense GRN mutations in 3 FTD patients from the Philippines were detected, but no known pathogenic mutations in other FTD-associated genes were found. In 45 subjects screened for C9orf72 repeat expansions, no pathogenic expansion (≥30 repeats) was identified, but there was a higher proportion of intermediate length (≥10-29 repeats) alleles in patients compared with controls (8/90 alleles, 8.9% vs. 9/164 alleles, 5.5%). Overall, we detected a mutation rate of 7.7% (4/52 patients) in our cohort. Given recent findings of enrichment of rare TREM2 variants (including R47C) in Alzheimer's disease, it is notable that we detected a homozygous TREM2 R47C carrier presenting with an FTD rather than an Alzheimer's disease phenotype.