Bp Deletion Research Articles

The Prevalence, Risk Factors, and Antimicrobial Resistance Determinants of Helicobacter pylori Detected in Dyspeptic Patients in North-Central Bangladesh.

Chronic infection of Helicobacter pylori represents a key factor in the etiology of gastrointestinal diseases, with high endemicity in South Asia. The present study aimed to determine the prevalence of H. pylori among dyspeptic patients in north-central Bangladesh (Mymensingh) and analyze risk factors of infection and antimicrobial resistance (AMR) determinants in the pathogen. Endoscopic gastrointestinal biopsy samples were collected from dyspeptic patients for a one-year period from March 2022 and were checked for the presence of H. pylori via the rapid urease test and PCR and further analyzed for the status of virulence factors vacA/cagA and genetic determinants related to AMR via PCR with direct sequencing or RFLP. Among a total of 221 samples collected, 80 (36%) were positive for H. pylori, with the vacA+/cagA+ genotype being detected in almost half of them. H. pylori was most prevalent in the age group of 41-50-year-olds, with it being more common in males and rural residents with a lower economic status and using nonfiltered water, though the rates of these factors were not significantly different from those of the H. pylori-negative group. Relatively higher frequency was noted for the A2147G mutation in 23S rRNA, related to clarithromycin resistance (18%, 7/39). Amino acid substitutions in PBP-1A (T556S) and GyrA (N87K and D91N) and a 200 bp deletion in rdxA were detected in samples from some patients with recurrence after treatment with amoxicillin, levofloxacin, and metronidazole, respectively. The present study describes the epidemiological features of H. pylori infection in the area outside the capital in Bangladesh, revealing the spread of AMR-associated mutations.

Bovine FRAS1: mRNA Expression Profile, Genetic Variations, and Significant Correlations with Ovarian Morphological Traits, Mature Follicle, and Corpus Luteum.

The amelioration of bovine fertility caused by a multi-factorial problem has always been a hot topic, among which the detection of available target genes is the most crucial. It was hypothesized that the Fraser extracellular matrix complex subunit 1 (FRAS1) gene detected by GWAS is involved in physiological activities such as ovarian development. Herein, unilateral ovaries from 2111 cows were used to examine the mRNA expression profile and polymorphisms of bovine FRAS1 and their associations with fertility-related characteristics. Firstly, it was confirmed that FRAS1 gene transcripts are expressed in various bovine tissues. Then, among five potential insertion-deletion (indel) loci, the 20 bp (named P3-D20-bp) and 15 bp (P4-D15-bp) deletion mutations were confirmed to be polymorphic with linkage equilibrium. Secondly, the P3-D20-bp polymorphism was significantly associated with ovarian weight and corpus luteum diameter in the metaestrus phase and ovarian length in the dioestrum stage. Additionally, both ovarian length and mature follicle diameter in metaestrus are significantly correlated with different genotypes of P4-D15-bp. Thirdly, the transcriptional expression of the FRAS1 gene in groups with a minimum value of ovarian weight or volume was significantly higher than the expression in groups with a maximum value. Instead of that, the more corpus luteum and mature follicles there are, the higher the transcription expression of the FRAS1 gene is. Furthermore, FRAS1 expression in cows with a heterozygous genotype (ID) of P3-D20-bp was significantly higher than others. Eventually, P3-D20-bp deletion could disturb the binding efficiency of WT1-I and Sox2 to FRAS1 sequence according to binding prediction, indicating that mutation may affect gene expression and traits by influencing the binding of transcription factors. Overall, the polymorphisms of P3-D20-bp and P4-D15-bp of the bovine FRAS1 gene significantly correlated to follicle or ovarian traits that could be applied in optimizing female fertility in cow MAS breeding programs.

Common mutation analysis of beta-thalassemia carriers by amplification refractory mutation system-polymerase chain reaction in cases diagnosed by capillary zone electrophoresis at National Public Health Laboratory, Nepal

Abstract BACKGROUND: Beta (β)-thalassemia is a globally prevalent genetic disorder with over 350 known mutations in the β-globin gene. Intervening-sequence (IVS) 1–5 (G→C), 619 bp deletion, IVS 1–1 (G →T), C-15 (G→A), frameshift (FS) 41/42 (−CTTT), FS 8/9 (+G), codon 19, A-G, and codon 17 (A→T) are some common mutations in β-thalassemia. This study aimed to identify and analyze the common mutation by amplification refractory mutation system polymerase chain reaction in β-thalassemia carriers diagnosed by capillary zone electrophoresis (CZE) at National Public Health Laboratory. MATERIALS AND METHODS: On the basis of archived data, the CZE performed on 781 samples between June 2019 and July 2022 was analyzed. Without regard to age, gender, race, or address, 121 samples were ultimately selected for mutation analysis during this time because they were identified as positive for the β-thalassemia traits based on high HbA2 levels of more than 3.5%. RESULTS: IVS 1–5 (G→C) was the most common mutation followed by 619 bp deletion. Five mutations namely IVS 1–5 (G→C), 619 bp deletion, FS 41/42 (−TTCT), FS 8/9 (+G), and codon 15 (G→A) accounted for 84.3% of cases. A single mutation was seen in 87 cases (71.9%), more than one mutation in 15 cases (12.4%) whereas 19 cases (15.7%) did not show any of the mutations tested. CONCLUSION: IVS 1–5 (G-C) is the most prevalent mutation in Southeast Asian nations, with the 619 bp deletion predominant in the Tharu community. Further research is needed to understand diverse ethnic groups and regions, enabling prenatal diagnostics and genetic counseling for β-thalassemia prevention.

Identification of two novel large deletions in FBN1 gene by next-generation sequencing and multiplex ligation-dependent probe amplification.

Mutations in fibrillin-1 (FBN1) are known to be associated with Marfan syndrome (MFS), an autosomal dominant connective tissue disorder. Most FBN1 mutations are missense or nonsense mutations. Traditional molecular genetic testing for the FBN1 gene, like Sanger sequencing, may miss disease-causing mutations in the gene's regulatory regions or non-coding sequences, as well as partial or complete gene deletions and duplications. Next-generation sequencing, multiplex ligation-dependent probe amplification and gap PCR were conducted on two MFS patients to screen for disease-causing mutations. We identified two large deletions in FBN1 from two MFS patients. One patient had a 0.23 Mb deletion (NC_000015.9:g.48550506_48779360del) including 5'UTR-exon6 of FBN1. The other patient harbored a 1416 bp deletion (NC_000015.9:g.48410869_48412284del) affecting the last exon, exon 66, of the FBN1 gene. Our results expanded the number of large FBN1 deletions and highlighted the importance of screening for large deletions in FBN1 in clinical genetic testing, especially for those with the classic MFS phenotype.

15-cis-Phytoene Desaturase and 15-cis-Phytoene Synthase Can Catalyze the Synthesis of β-Carotene and Influence the Color of Apricot Pulp.

Fruit color affects its commercial value. β-carotene is the pigment that provides color for many fruits and vegetables. However, the molecular mechanism of β-carotene metabolism during apricot ripening is largely unknown. Here, we investigated whether β-carotene content affects apricot fruit color. First, the differences in β-carotene content between orange apricot 'JTY' and white apricot 'X15' during nine developmental stages (S1-S9) were compared. β-carotene contents highly significantly differed between 'JTY' and 'X15' from S5 (color transition stage) onwards. Whole-transcriptome analysis showed that the β-carotene synthesis genes 15-cis-phytoene desaturase (PaPDS) and 15-cis-phytoene synthase (PaPSY) significantly differed between the two cultivars during the color transition stage. There was a 5 bp deletion in exon 11 of PaPDS in 'X15', which led to early termination of amino acid translation. Gene overexpression and virus-induced silencing analysis showed that truncated PaPDS disrupted the β-carotene biosynthesis pathway in apricot pulp, resulting in decreased β-carotene content and a white phenotype. Furthermore, virus-induced silencing analysis showed that PaPSY was also a key gene in β-carotene biosynthesis. These findings provide new insights into the molecular regulation of apricot carotenoids and provide a theoretical reference for breeding new cultivars of apricot.

Gene Mapping of a Yellow-to-Lethal Mutation Based on Bulked-Segregant Analysis-Seq in Soybean

Plant photosynthesis is mainly dependent on leaf color, and this has an impact on yield. Mutants lacking in chlorophyll have been analyzed to gain insight into the genetic processes involved in photosynthesis, chloroplast development, and chlorophyll metabolism. A yellow-to-lethal mutant, ytl, was selected from the M6 generation of the 60Coγ ray irradiation-treated soybean cultivar Nannong 1138-2. The mutant exhibited reduced chlorophyll content, with the thylakoid structure disrupted. Segregation of the cross between Williams 82 (W82) and ytl indicated that a recessive allele controlled yellow-to-lethal traits. The bulked-segregant analysis (BSA)-Seq method performed preliminary mapping, followed by simple sequence repeat (SSR) marker validation and further mapping. The candidate gene was mapped to a 418 Kb region containing 53 genes. High-throughput sequencing and first-generation sequencing results showed a two bp deletion in the second exon of Glyma.08g106500, leading to a frameshift mutation in ytl. As a promising candidate gene, Glyma.08g106500 encoded a chloroplast-localized pentatricopeptide repeat (PPR) domain-containing protein involved in the assembly of chloroplast proteins. These results will contribute to cloning the mutant ytl gene and provide insight into the regulatory processes controlling photosynthesis and chloroplast development and growth in soybean.

Kagami Ogata syndrome: a small deletion refines critical region for imprinting

Kagami–Ogata syndrome is a rare imprinting disorder and its phenotypic overlap with multiple different etiologies hampers diagnosis. Genetic etiologies include paternal uniparental isodisomy (upd(14)pat), maternal allele deletions of differentially methylated regions (DMR) in 14q32.2 or pure primary epimutations. We report a patient with Kagami–Ogata syndrome and an atypical diagnostic odyssey with several negative standard-of-care genetic tests followed by epigenetic testing using methylation microarray and a targeted analysis of whole-genome sequencing to reveal a 203 bp deletion involving the MEG3 transcript and MEG3:TSS-DMR. Long-read sequencing enabled the simultaneous detection of the deletion, phasing, and biallelic hypermethylation of the MEG3:TSS-DMR region in a single assay. This case highlights the challenges in the sequential genetic testing paradigm, the utility of long-read sequencing as a single comprehensive diagnostic assay, and the smallest reported deletion causing Kagami–Ogata syndrome allowing important insights into the mechanism of imprinting effects at this locus.

Genetic analysis of a Chinese pedigree with an allele dropout at the HLA-B locus

To delineate a deletional mutation of the HLA-B gene in a Chinese pedigree. A female patient with acute myeloid leukemia who had visited Liuzhou People's Hospital in April 2022 was selected as the study subject. Routine human leukocyte antigen (HLA) was determined by using PCR-sequence specific oligonucleotide polymorphism (PCR-SSOP) and PCR-sequence-based typing (PCR-SBT) methods. Next generation sequencing (NGS) was used to validate the candidate variant in the HLA-B gene. The PCR-SBT and SSOP results for the HLA-B locus were inconsistent for the patient and her daughter. The SSOP results of the two individuals were HLA-B*35:01, 40:02 and HLA-B*35:01, 40:01, respectively. However, the PCR-SBT results has indicated a mismatch with the nearest HLA-B*35:01 at exon 4. NGS results showed that the HLA-B*35:01 had a 9 bp deletion in the intron 5. The patient's husband was HLA-B*40:01, 58:01, which was normal. The variant in intron 5 of the HLA-B gene in this pedigree has mapped to a primer-binding region for the SBT reagent, which has affected the accuracy of PCR-SBT results.

Knockout of a rice K5.2 gene increases Ca accumulation in the grain.

Rice is a staple food for half of the world's population, but it is a poor dietary source of calcium (Ca) due to the low concentration. It is an important issue to boost Ca concentration in this grain to improve Ca deficiency risk, but the mechanisms underlying Ca accumulation are poorly understood. Here, we obtained a rice (Oryza sativa) mutant with high shoot Ca accumulation. The mutant exhibited 26%-53% higher Ca in shoots than did wild-type rice (WT) at different Ca supplies. Ca concentration in the xylem sap was 36% higher in the mutant than in the WT. There was no difference in agronomic traits between the WT and mutant, but the mutant showed 25% higher Ca in the polished grain compared with the WT. Map-based cloning combined with a complementation test revealed that the mutant phenotype was caused by an 18-bp deletion of a gene, OsK5.2, belonging to the Shaker-like K+ channel family. OsK5.2 was highly expressed in the mature region of the roots and its expression in the roots was not affected by Ca levels, but upregulated by low K. Immunostaining showed that OsK5.2 was mainly expressed in the pericycle of the roots. Taken together, our results revealed a novel role for OsK5.2 in Ca translocation in rice, and will be a good target for Ca biofortification in rice.

Mitochondrial DNA editing in potato through mitoTALEN and mitoTALECD: molecular characterization and stability of editing events

BackgroundThe aim of this study was to evaluate and characterize the mutations induced by two TALE-based approaches, double-strand break (DSB) induction by the FokI nuclease (mitoTALEN) and targeted base editing by the DddA cytidine deaminase (mitoTALECD), to edit, for the first time, the mitochondrial genome of potato, a vegetatively propagated crop. The two methods were used to knock out the same mitochondrial target sequence (orf125).ResultsTargeted chondriome deletions of different sizes (236–1066 bp) were induced by mitoTALEN due to DSB repair through ectopic homologous recombination of short direct repeats (11–12 bp) present in the target region. Furthermore, in one case, the induced DSB and subsequent repair resulted in the amplification of an already present substoichiometric molecule showing a 4288 bp deletion spanning the target sequence. With the mitoTALECD approach, both nonsense and missense mutations could be induced by base substitution. The deletions and single nucleotide mutations were either homoplasmic or heteroplasmic. The former were stably inherited in vegetative offspring.ConclusionsBoth editing approaches allowed us to obtain plants with precisely modified mitochondrial genomes at high frequency. The use of the same plant genotype and mtDNA region allowed us to compare the two methods for efficiency, accuracy, type of modifications induced and stability after vegetative propagation.

Allelic variation of BBX24 is a dominant determinant controlling red coloration and dwarfism in pear.

Variation in anthocyanin biosynthesis in pear fruit provides genetic germplasm resources for breeding, while dwarfing is an important agronomic trait, which is beneficial to reduce the management costs and allow for the implementation of high-density cultivation. Here, we combined bulked segregant analysis (BSA), quantitative trait loci (QTL), and structural variation (SV) analysis to identify a 14-bp deletion which caused a frame shift mutation and resulted in thepremature translation termination of a B-box (BBX) family of zinc transcription factor, PyBBX24, and its allelic variation termed PyBBX24ΔN14. PyBBX24ΔN14 overexpression promotes anthocyanin biosynthesis in pear, strawberry, Arabidopsis, tobacco, and tomato, while that of PyBBX24 did not. PyBBX24ΔN14 directly activates the transcription of PyUFGT and PyMYB10 through interaction with PyHY5. Moreover, stable overexpression of PyBBX24ΔN14 exhibits a dwarfing phenotype in Arabidopsis, tobacco, and tomato plants. PyBBX24ΔN14 can activate the expression of PyGA2ox8 via directly binding to its promoter, thereby deactivating bioactive GAsand reducing the plant height. However, the nuclear localization signal (NLS) and Valine-Proline (VP) motifs in the C-terminus of PyBBX24 reverse these effects. Interestingly, mutations leading to premature termination of PyBBX24 were also identified in red sports of un-related European pear varieties. We conclude that mutations in PyBBX24 gene link both an increase in pigmentation and a decrease in plantheight.

Distribution of Epstein-Barr virus in the oral cavity of Thais with various oral mucosal conditions

ObjectivesWe aimed to examine the presence of EBV, EBV strains, and variants among 3 oral conditions including normal oral mucosa (NOM), oral potentially malignant disorders/oral cancer (OPMDs/OC) and non-OPMDs/OC in a group of Thais. Material and methodsOral exfoliated cells were obtained from 315 participants living in the northeastern and central regions of Thailand. The participants were divided into 3 groups encompassing the NOM, the OPMDs/OC and the non-OPMDs/OC groups. The presence of EBV was first determined by PCR using primers for LMP1 gene. Subsequently, EBV strains of EBNA3c and variants based on LMP1 sequences were determined by real-time PCR. ResultsThe prevalence of EBV in OPMDs/OC, non-OPMDs/OC and NOM were 72.0 %, 56.2 %, and 27.2 % respectively. EBV type A, B and AB were found in 52.1 %, 32.1 % and 15.8 % of all positive samples, respectively. The percentage of participants with EBV type A was more prominent in the NOM group (72.0 %) compared to the non-OPMDs/OC (54.8 %) and the OPMDs/OC group (41.8 %) whereas EBV type B was higher in the OPMDs/OC group (35.8 %) compared to the non-OPMDs/OC (31.5 %) and the NOM (24.0 %) groups. Regarding EBV variants, 30-bp deletion LMP1 variant (del-LMP1) which is more associated with malignant transformation was predominately found in the OPMDs/OC (32.8 %) and the non-OPMDs/OC (38.4 %) groups compared to the NOM group (20.0 %). ConclusionsHigh frequency of EBV was demonstrated in the OPMDs/OC group. EBV type A was more predominant in the NOM group whereas EBV type B was more prevalent in the OPMDs/OC group. The del-LMP1 variant was more common in the OPMDs/OC and the non-OPMDs/OC groups.

Genetic variations at BBX24 and MYB110a loci regulated anthocyanin accumulation in pear bud sports

Genetic variation contributes to the phenotypic diversity of plants. Most red pear cultivars that accumulate anthocyanins originated from somatic variation and are termed bud sports. ‘Hongzaosu’ is a bud sport derived from the green pear ‘Zaosu’. A new genetic map was constructed from a population derived from the cross of ‘Yuluxiang’ and ‘Hongzaosu’, from which PpBBX24 was identified as a crucial gene controlling anthocyanin accumulation. Genetic and phylogenetic evidence revealed that PpBBX24 is a repressor of anthocyanin biosynthesis in pear. A 14 bp deletion was detected in the third exon of PpBBX24 in ‘Hongzaosu’, resulting in premature termination of protein translation. The truncated PpBBX24 protein was a positive regulator of anthocyanin biosynthesis by activating the transcription of PpCHS and PpUFGT. Three independent variations in the BBX24 coding region that led to premature termination of translation were detected in other pear bud sports and the progeny of a bud sport that all accumulate anthocyanins. The genome of ‘Hongzaosu’ was assembled using both long-read and short-read sequences. By combining genomic and transcriptomic data, allele-specific expression of PpMYB110a was observed in the fruit skin of ‘Hongzaosu’, which was likely caused by large variation in the promoter. This work enhances understanding of coloration in red pears and provides a novel genomic resource for further studies on ‘Hongzaosu’ pear.

Specific Differences in AMELX and AMELY Genes from Sumatran Tigers, Panthera tigris sumatrae (Felidae), for Molecular Sex Identification

Sex determination by DNA-based molecular techniques in Sumatran tigers needs to be investigated and developed for forensic and population genetic purposes. The amelogenin gene is a marker commonly used for sex determination. In some species, the difference between the AMELX and AMELY sequences has been reported to be in the intron region. However, the difference between the AMELX and AMELY sequences in the Sumatran tiger is unknown. Therefore, it is necessary to investigate the sequence differences in introns between AMELX and AMELY Sumatran tigers to determine the specific differences between male and female samples. This study aimed to analyse the sequence of nucleotide bases in the Sumatran tiger amelogenin gene introns based on the nucleotide base sequences in the amelogenin gene introns. The method in this research is descriptive, with a molecular observation of the AMELX and AMELY Sumatran tiger sequences. The amplified samples were sequenced, and it was found that the lengths of the AMELX and AMELY Sumatran tiger sequences were 215 bp and 194 bp, respectively, with a 21 bp deletion in the AMELY sequence.

A novel 11 base pair deletion in KMT2C resulting in Kleefstra syndrome 2.

Haploinsufficiency of the Lysine Methyltransferase 2C (KMT2C) gene results in the autosomal dominant disorder, Kleefstra syndrome 2. It is an extremely rare neurodevelopmental condition, with 14 previous reports describing varied clinical manifestations including dysmorphic features, delayed psychomotor development and delayed growth. Here, we describe a female with global developmental delay, attention deficit disorder, dyspraxia, short stature and subtle non-specific dysmorphic features. To identify causative mutations, whole exome sequencing was performed on the proband and her younger brother with discrete clinical presentation. Whole exome sequencing identified a novel de novo heterozygous 11 bp deletion in KMT2C (c.1759_1769del), resulting in a frameshift mutation and early termination of the protein (p.Gln587SerfsTer7). This variant is the second-most N-terminal reported mutation, located 4171 amino acids upstream of the critical enzymatically active SET domain (required for chromatin modification and histone methylation). The majority of the other reported mutations are frameshift mutations upstream of the SET domain and are predicted to result in protein truncation. It is thought that truncation of the SET domain, results functionally in an inability to modify chromatin through histone methylation. This report expands the clinical and genetic characterisation of Kleefstra syndrome 2.

TAF7L regulates early stages of male germ cell development in the rat.

Male germ cell development is dependent on the orchestrated regulation of gene networks. TATA-box binding protein associated factors (TAFs) facilitate interactions of TATA-binding protein with the TATA element, which is known to coordinate gene transcription during organogenesis. TAF7 like (Taf7l) is situated on the X chromosome and has been implicated in testis development. We examined the biology of TAF7L in testis development using the rat. Taf7l was prominently expressed in preleptotene to leptotene spermatocytes. To study the impact of TAF7L on the testis we generated a global loss-of-function rat model using CRISPR/Cas9 genome editing. Exon 3 of the Taf7l gene was targeted. A founder was generated possessing a 110 bp deletion within the Taf7l locus, which resulted in a frameshift and the premature appearance of a stop codon. The mutation was effectively transmitted through the germline. Deficits in TAF7L did not adversely affect pregnancy or postnatal survival. However, the Taf7l disruption resulted in male infertility due to compromised testis development and failed sperm production. Mutant germ cells suffer meiotic arrest at late zygotene/early pachynema stages, with defects in sex body formation. This testis phenotype was more pronounced than previously described for the subfertile Taf7l null mouse. We conclude that TAF7L is essential for male germ cell development in the rat.

White and other fur colourations and hybridization in golden jackals (Canis aureus) in the Carpathian basin

The golden jackal (Canis aureus) is a reoccurring species in the centre of the Carpathian basin, in Hungary. In total, 31 golden jackal tissue samples were collected, from 8 white-coated, 2 black-coated and one mottled animal across Hungary. Sequences and fragment length polymorphisms were studied for white colour (MC1R), and for black coat colouration (CBD103). In each white animal, the most widespread mutation causing white fur colour in dogs in homozygous form was detected. Three animals were found to carry the mutation in heterozygous form. The two black golden jackals were heterozygous for the 3 bp deletion in CBD103 that mutation for black coat colouration in dogs, and one of them also carried the mutation causing white fur. None of the white animals showed signs of hybridization, but both the black and the mottled coloured individuals were found to be hybrids based on genetic testing. Kinship was found three times, twice between white animals, and once between a white animal and an agouti animal carrying the mutation of white coat. Our results confirm the findings that golden jackal–dog hybrids may occur without human intervention, and the detected mutation causing white fur colour in golden jackals could possibly be due to an early hybridization event.

Development of the Cas12a-based microdeletion and microinsertion detection system

CRISPR/Cas-based systems are widely used as genome editing systems, nucleic acid detection systems and molecular visualization instruments [1]. In our laboratory using CRISPR/Cas9 technology we have obtained several KO mouse lines harboring deletions ranging from 2 to 20 base pairs. While 20 bp deletions are easily PCR-detected, when it comes to 2 bp deletions it is essential to genotype numerous mice using time-consuming Sanger sequencing. We propose a microdeletion/microinsertion detection system based on Cas12a nuclease fromLachnospiraceaebacterium (LbCas12a). Its active complex consists of the Cas12a enzyme and one crisprRNA [2]. A special sequence called protospacer adjacent motif (PAM) is required for target recognition by LbCas12a. In our laboratory we have discovered new PAM TTAA recognized by LbCas12a [3]. Via agarose electrophoresis and fluorescent analysis using FAM-labeled probes we have shown that new PAM allowed detection of 1 bp substitutions in target DNAin vitro. Also we have tested different FAM-labeled probes and have shown that AT-rich probes longer than 10 bp are cleaved most effectively. Finally we have used our system for detecting 2 bp deletions in Pde6b-KO mice and Grin3A-KO mice and successfully distinguished these mice from wild type mice.
 In conclusion, new PAM TTAA greatly increases specificity of DNA cleavage allowing to use this system as an instrument for rapid detection if microdeletions in mice.
 This work was supported by a Saint Petersburg State University grant for the development of scientific research (ID 94030690) and the Genome Research Centre development program “Kurchatov Genome Centre–PNPI” (agreement No. 075-15-2019-1663)

A naturally occurring 31 bp deletion in TEOSINTE BRANCHED1 causes branched ears in maize

The female inflorescence, or ear, of maize develops no branch meristem (BM), which differs from the male inflorescence, or tassel. While the mutations of some well documented genes, such as fea2/3/4 and ramosa1/2/3, can cause the branched architecture of ears in maize, such mutations also change the normal phenotypic performance of the tassels. In the present study, a natural maize mutant with branched ears, named branched ear1 (be1), was characterized. be1 shows several branched ears at the base of the central ear with unchanged architecture of the tassels. Besides, both the branched and central ears of be1 possess regularly arranged kerels. The phenotypic characteristics of be1 differ completely from those reported mutants of fasciated ears or RAMOSA-like ears in maize. An SEM survey at the very early development stage showed that meristems with three protrusions, similar to the BM in tassels, were present during the development of the branched ears in be1. Gene mapping and sequence alignment suggested that TEOSINTE BRANCHED1 (TB1) was the candidate gene of BE1. Further verification showed that a be1-specific 31 bp deletion at the downstream of BE1 led to statistically reduced expression of this gene in the immature ear, which serves as the potential causal reason for the branched ears of be1. CRISPR/Cas9-based gene editing downstream of TB1 complemented the phenotypic architecture of branched ears, suggesting that TB1 was the target of BE1, and it was named as ZmTB1be1. The results of the present study implied a novel function of TB1 in female inflorescence development, rather than shaping the plant architecture in maize. Meanwhile, further functional dissection of ZmTB1be1 might shed new light on TB1, the most famous domestication related gene in maize.

Novel characterization of Alzheimer’s disease genetic loci using long‐read sequencing

AbstractBackgroundAlzheimer’s disease (AD) is up to 60‐80% heritable, but less than ∼20% is explained by studies analyzing single nucleotide variants (SNVs). One limitation of short‐read whole‐genome sequencing (SRS) is the standard read length of 150 base pairs, which does not enable the detection of longer structural variants (SVs). Here we sequenced individuals using long‐read sequencing (LRS), which can sequence reads with an average length of ∼20 kilobases, allowing us to identify SVs previously uncaptured.MethodAll participants underwent whole‐genome LRS (∼15x coverage) and a subgroup (84%) underwent SRS. Out of 576 participants (47% males, age = 70.6±7.9 y.o.), 115 were diagnosed with AD or mild cognitive impairment, 365 were healthy controls, and 96 were diagnosed with a synucleinopathy (either Parkinson or Lewy Body disease). Eighty‐three index SNVs from loci associated with AD risk through GWAS (Bellenguez et al., Nature Genetics 2022) were genotyped with 30x coverage SRS, in addition to APOE2‐4. A 1Mbp window was defined around these SNVs to construct the discovery range for SVs (Sniffles2 population mode). Linkage disequilibrium (LD) was assessed between SNVs and SVs (CubeX).ResultA total of 14854 SVs were found across the AD risk loci (Figure1). After LD calculation, N = 197 SVs had a R2>0.1 (Figure2). The SVs with the highest LD (R2>0.7) are reported in Table1, among which there is a 322 bp deletion in the 3’ UTR region of the TMEM106B in high LD (R2 = 0.918) with the intronic SNV rs13237518. This TMEM106B locus has been previously associated with the risk of frontotemporal lobar dementia with TDP‐43 pathology in addition to AD, but the causative SNV has not been identified yet. This large deletion may mediate TMEM106B’s risk‐modulating role in AD and FTLD‐TDP (Chemparathy et al. medRxiv 2023). At the complex MAPT locus, three SVs showed a high LD with rs199515.ConclusionInsights into the role of SVs in neurodegenerative disorders have been hampered due to limitations with SRS. Using LRS in a large AD‐related sample, we characterized for the first time the genetic variation of SVs in known AD risk loci and provide a roadmap to identify potential causal SVs driving the AD association signal.