Insertion Deletion Research Articles

Characterization of sequences, expression profiling, and natural allelic variation analysis of the MYC gene family in sorghum (Sorghum bicolor)

Sorghum aphid (Melanaphis sacchari) and head smut fungi (Sporisorium reilianum) infesting sorghum cause delayed growth and development, and reduce yield and quality. This study use bioinformatics and molecular biological approaches to profile the gene expression pattern during sorghum development and under pest infestation, and analyzed the natural allelic DNA variation of sorghum MYC gene family. The findings provide insights for potential application in breeding the stress resistant and high productivity sorghum varieties. The results indicated that there are 28 MYC genes identified in sorghum genome, distributed on 10 chromosomes. The bHLH_MYC_N and HLH domains are the conserved domains of the MYC gene in sorghum. Gene expression analysis showed that SbbHLH35.7g exhibited high expression levels in leaves, SbAbaIn showed strong expression in early grains, and SbMYC2.1g showed high expression levels in mature pollen. In anti-aphid strains at the 5-leaf stage, SbAbaIn, SbLHW.4g and SbLHW.2g were significantly induced in leaves, while SbbHLH35.7g displayed the highest expression level in panicle tissue, which was significantly induced by the infection of head smut. Promoter cis-element analysis identified methyl jasmonate (MJ), abscisic acid (ABA), salicylic acid (SA) and MYB-binding sites related to drought-stress inducibility. Furthermore, genomic resequencing data analysis revealed natural allelic DNA variations such as single nucleotide polymorphism (SNP) and insertion-deletion (INDEL) for the key SbMYCs. Protein interaction network analysis using STRING indicated that SbAbaIn interacts with TIFYdomain protein, and SbbHLH35.7g interacts with MDR and imporin. SbMYCs exhibited temporal and spatial expression patterns and played vital roles during the sorghum development. Infestation by sugarcane aphids and head smut fungi induced the expression of SbAbaIn and SbbHLH35.7g, respectively. SbAbaIn modulated the jasmonic acid (JA) pathway to regulate the expression of defensive genes, conferring resistance to insects. On the other hand, SbbHLH35.7g participated in detoxification reactions to defend against pathogens.

CRISPR/Cas9 targeting of passenger single nucleotide variants in haploinsufficient or essential genes expands cancer therapy prospects

CRISPR/Cas9 technology has effectively targeted cancer-specific oncogenic hotspot mutations or insertion–deletions. However, their limited prevalence in tumors restricts their application. We propose a novel approach targeting passenger single nucleotide variants (SNVs) in haploinsufficient or essential genes to broaden therapeutic options. By disrupting haploinsufficient or essential genes through the cleavage of DNA in the SNV region using CRISPR/Cas9, we achieved the selective elimination of cancer cells without affecting normal cells. We found that, on average, 44.8% of solid cancer patients are eligible for our approach, a substantial increase compared to the 14.4% of patients with CRISPR/Cas9-applicable oncogenic hotspot mutations. Through in vitro and in vivo experiments, we validated our strategy by targeting a passenger mutation in the essential ribosomal gene RRP9 and haploinsufficient gene SMG6. This demonstrates the potential of our strategy to selectively eliminate cancer cells and expand therapeutic opportunities.

Exploring the potential of data augmentation in poetry generation with small-scale corpora

Poetry generation is a complex task in the field of natural language processing, especially when working with small datasets. Data augmentation techniques have been shown to be an effective way to improve the performance of deep learning models in various tasks, including image classification and speech recognition. Therefore, this study focuses on exploring the impact of four different data augmentation methods - Synonym Replacement, Random Insertion, Random Swap, and Random Deletion - on the performance of poetry generation with a small poetry dataset. The results of the study reveal that Random Insertion performed well in terms of Bilingual Evaluation Understudy (BLEU), Recall-Oriented Understudy for Gisting Evaluation (ROUGE), and manual evaluation when compared to other data augmentation techniques. Synonym Replacement performed poorly in all three evaluations. This study confirms the potential value of data augmentation technology in poetry generation tasks and provides innovative perspectives and directions for future research in this area. Data augmentation can be employed to help address the problem of limited data in poetry generation tasks and enhance the efficiency of deep learning models. Future research could focus on exploring more advanced data augmentation techniques and their impact on poetry generation tasks.

Abstract 5023: Analytical validation of the Oncomine™ Dx Express Test using liquid biopsy samples as an IUO assay in a CAP/CLIA lab

Abstract Introduction: The Oncomine™ Dx Express Test (ODxET) is a next-generation sequencing (NGS) test. It can be used as an investigational use only (IUO**) test to detect somatic DNA and RNA alterations in human cell-free total nucleic acid (cfTNA) isolated from plasma samples. This study outlines the analytical validation of the ODxET for relevant single nucleotide variants (SNV) and insertion/deletions (INDEL) in liquid biopsy specimens. The Ion Torrent Genexus™ platform has a short turnaround time with minimal operator input. This study was performed in a College of American Pathologists (CAP™) accredited and Clinical Laboratory Improvement Act (CLIA) licensed laboratory. Methods: Forty-seven unique samples (39 clinical, 3 derived, 2 reference and 3 commercially available DNA controls) were analyzed. All experiments began with sample extraction, except the lower input limit of detection (LLOD). We evaluated limit of detection, variant calling accuracy, and precision. Two control NIST™ samples were sequenced with 3 technical replicates each for INDEL and SNVs analytical accuracy. The LOD was evaluated at both the target input of 2ng/µl (40ng), and the minimum input,0.33ng/µl (6.6ng) with allele frequency (AF) levels at 1%, 0.5%, and 0.25%. The 39 clinical plasma samples were used for variant calling accuracy. Inter-assay reproducibility was assessed by sequencing DNA controls across four runs by different operators over multiple days/systems. Intra-assay repeatability was assessed by sequencing 4 samples in replicate on the same run by one operator on the same day/system. Results: The analytical accuracy and NPV was 100% for SNV/INDEL variants. The 40ng input LOD sensitivity was 96% for SNVs at 0.5% AF and 100% for INDELs at 0.25% Minimum AF. The lower input LOD sensitivity was 89% for SNVs and 100% for INDEL at 1% AF. Variant calling accuracy on samples with pre-characterized data was 100% for the 13 positive samples. Specificity was >99% for all samples including both healthy donor and positive samples. Repeatability was 100% for INDELs and >96% for SNVs, reproducibility was 100% for both variant classes. Conclusion: This validation study to estimate analytical accuracy, limit of detection, variant calling accuracy and precision has demonstrated that the ODxET assay is highly robust in detecting SNV and INDEL mutations in plasma samples and is suitable for use in a CAP/CLIA laboratory as an IUO assay.**=In Vitro Diagnostic use only. Not available in all regions including North America. Citation Format: Juan C. Espinoza, Rajendra Ramsamooj, Akemi Yuki, John Williamson, Diarra Hassell, Madhu Jasti, Suzanne Salazar, David Ginzinger. Analytical validation of the Oncomine™ Dx Express Test using liquid biopsy samples as an IUO assay in a CAP/CLIA lab [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5023.

Abstract 4402: A genomic insertion/deletion variant activates immune escape and tumor promoting programs in breast cancer

Abstract Purpose: Despite all the knowledge we have concerning cancer cells’ immunesuppressive ability, we still don’t understand whether the mechanisms upregulatingimmune suppression in cancer cells are patient specific. This information is critical forthe generation of tailored treatments that recruit the immune system for the eradicationof cancer disease. In the past, we showed that super-enhancers, composed of cluster ofenhancers spanning over long genomic regions characterized by having potent generegulatory activity, are critical for the upregulation of immune suppressive genes,including CD47, within the tumor. Recently we found a DNA insertion/deletion variant (8bp in size) within a super-enhancer located between the CD47 and Linc00636 genes,that is present more often in European (53%) than Asian (12%) populations and whichassociates with breast cancer. Moreover, sequence analyses of patient-derived breasttumors (44) and gynecological tumor samples (23) demonstrated a significantassociation of this insert with breast cancer and not with gynecological cancer. Thus, wehypothesize that the insertion variant regulates genes that drive immune escape andbreast cancer specifically in patient populations that carry the insert version. Methods: To test this, we CRISPR deleted the variant’s insertion and CRISPR-VP64induced its activation and quantified its effect on target genes and encoded proteinsusing high throughput RNA sequencing and immunofluorescence flow cytometryanalyses respectively. Results: CRISPR-VP64 induction of the 8bp insertion confirmed that the insertion issufficient to regulate gene expression of CD47 and of an anti-tumor, long intergenicnon-coding RNA (Linc00636) in breast cancer cells. Furthermore, we observed thatCRISPR deletion of the 8bp insertion increases CD47 levels, while dramaticallyincreasing expression of Linc00636 and reducing the expression of CD44, CXCL14,MUC1 genes involved in invasion and migration. Our work shows that the insertion ofan 8bp is responsible for stabilizing upregulation of CD47 (an immune suppressivesignal), downregulating Linc00636 (a modulator of tumor metastasis), and upregulating,as a result, the expression of cell surface receptors involved in metastasis andtumorigenesis. Conclusions: Here we found a genomic insert within a super-enhancer that unleashesthe upregulation of Linc00636 and of a program that promotes breast cancer, mostlikely in populations of patients carrying this variant. Our findings are important tounderstand patient specific genomic bifunctional modulators of immune suppressionand tumor progression for the design of more personalized treatments geared towardprecision medicine. Citation Format: Paola Betancur, Carolina DiBenedetto, Daniza Diane Acenas, Anthony Rodriguez, Alysia Thach. A genomic insertion/deletion variant activates immune escape and tumor promoting programs in breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4402.

Abstract 3886: Analysis of clonal heterogeneity within paired primary and metastatic tumor samples of patients with solid tumors and implications for neoantigen-based personalized cancer vaccines

Abstract Background: Neoantigen-based personalized cancer vaccines carry significant promise in treating solid malignancies. However, there are uncertainties regarding the choice between the primary or the metastatic tumor for neoantigen prediction in individual patients. Here, we conducted a thorough examination of somatic variations in 676 patients who had paired primary and metastatic solid tumors. Methods: Patients were enrolled in the Total Cancer Care protocol (NCT03977402) to which patients provided an IRB-approved written informed consent within the Oncology Research Information Exchange Network (ORIEN). Whole-exome sequencing of 756 primary and metastatic tumor pairs was performed (N = 676 patients). These included Genitourinary (n=83), Gynecological (n=97), Gastrointestinal (n=213), Thoracic (n=33), Cutaneous (n=24), Breast (n=108), Endometrial (n=49), Sarcoma (n=35), Head-and-Neck (n=106) and others (n=8). The data was analyzed through the ORIEN AVATAR Molecular Analysis Pipeline for somatic mutation variant detection and variant annotation. In this analysis, we focused on somatic events that result in an in-frame alteration (such as missense, in-frame deletion and in-frame insertion) and out-of-frame protein-altering mutations (such as frameshifts, de novo start, out-of-frame, and nonstop gain). Clonal population structure was determined based on pyclone-vi. Results: For in-frame events, bladder cancer, melanoma, and gynecological cancers shared close to 75% of the mutations between paired primary and metastatic cases. In contrast, sarcoma and thyroid cancer had a low overlap (~ 25%) of variants. For out-of-frame events, these events tend to have a lower proportion of shared somatic variants between primary and metastasis than in-frame variants. Oncogenic drivers (e.g., BRAF V600E, KRAS G12A, and TP53 loss-of-function) were highly likely to be present in both paired primary and metastatic tumors. Next, we performed additional analysis on evolutionary selection of protein-coding variants via dN/dS calculation. We found no significant global shift in dN/dS ratio between paired primary and metastatic tumors across malignancies. However, we found increased selection of protein-coding variants in brain and liver metastatic sites, which correlated with increased homologous recombination deficiency within these sites. Conclusions: Our analysis demonstrates genetic variations that exist when comparing paired primary and metastatic tumors that appear to vary by histology. Variants are potentially undergoing negative selection supported by the preferential loss of out-of-frame events in metastatic tumors and positive selection in specific metastatic sites. Overall, understanding the clonal structure will be key to neoantigen prediction for effective neoantigen-based vaccines. Citation Format: Alyssa Obermayer, Timothy Shaw, Darwin Chang, Joshua Davis, Jamie K. Teer, Xiaoqing Yu, Xuefeng Wang, Dale Hedges, Aik Choon Tan, Robert Rounbehler, Abdul Rafeh Naqash, Margaret Gatti-Mays, Aakrosh Ratan, Martin McCarter, Howard Colman, Igor Puzanov, Susanne Arnold, Michelle Churchman, Patrick Hwu, William Dalton, George Weiner, Jose Conejo-Garcia, Paulo C. Rodriguez, Bodour Salhia, Ahmad A. Tarhini. Analysis of clonal heterogeneity within paired primary and metastatic tumor samples of patients with solid tumors and implications for neoantigen-based personalized cancer vaccines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3886.

Identification of pleiotropic loci mediating structural and non-structural carbohydrate accumulation within the sorghum bioenergy association panel using high-throughput markers.

Molecular characterization of diverse germplasm can contribute to breeding programs by increasing genetic gain for sorghum [Sorghum bicolor (L.) Moench] improvement. Identifying novel marker-trait associations and candidate genes enriches the existing genomic resources and can improve bioenergy-related traits using genomic-assisted breeding. In the current scenario, identifying the genetic loci underlying biomass and carbon partitioning is vital for ongoing efforts to maximize each carbon sink's yield for bioenergy production. Here, we have processed a high-density genomic marker (22 466 550) data based on whole-genome sequencing (WGS) using a set of 365 accessions from the bioenergy association panel (BAP), which includes ~19.7 million (19 744 726) single nucleotide polymorphism (SNPs) and 2.7 million (~2 721 824) insertion deletions (indels). A set of high-quality filtered SNP (~5.48 million) derived markers facilitated the assessment of population structure, genetic diversity, and genome-wide association studies (GWAS) for various traits related to biomass and its composition using the BAP. The phenotypic traits for GWAS included seed color (SC), plant height (PH), days to harvest (DTH), fresh weight (FW), dry weight (DW), brix content % (BRX), neutral detergent fiber (NDF), acid detergent fiber (ADF), non-fibrous carbohydrate (NFC), and lignin content. Several novel loci and candidate genes were identified for bioenergy-related traits, and some well-characterized genes for plant height (Dw1 and Dw2) and the YELLOW SEED1 locus (Y1) were validated. We further performed a multi-variate adaptive shrinkage analysis to identify pleiotropic QTL, which resulted in several shared marker-trait associations among bioenergy and compositional traits. Significant marker-trait associations with pleiotropic effects can be used to develop molecular markers for trait improvement using a marker-assisted breeding approach. Significant nucleotide diversity and heterozygosity were observed between photoperiod-sensitive and insensitive individuals of the panel. This diverse bioenergy panel with genomic resources will provide an excellent opportunity for further genetic studies, including selecting parental lines for superior hybrid development to improve biomass-related traits in sorghum.

Development of a novel five dye insertion/deletion (INDEL) panel for ancestry determination.

The use of genetic markers, specifically Short Tandem Repeats (STRs), has been a valuable tool for identifying persons of interest. However, the ability to analyze additional markers including Single Nucleotide Polymorphisms (SNPs) and Insertion/Deletion (INDELs) polymorphisms allows laboratories to explore other investigative leads. INDELs were chosen in this study because large panels can be differentiated by size, allowing them to be genotyped by capillary electrophoresis. Moreover, these markers do not produce stutter and are smaller in size than STRs, facilitating the recovery of genetic information from degraded samples. The INDEL Ancestry Informative Markers (AIMs) in this study were selected from the 1000 Genomes Project based on a fixation index (FST) greater than 0.50, high allele frequency divergence, and genetic distance. A total of 25 INDEL-AIMs were optimized and validated according to SWGDAM guidelines in a five-dye multiplex. To validate the panel, genotyping was performed on 155 unrelated individuals from four ancestral groups (Caucasian, African, Hispanic, and East Asian). Bayesian clustering and principal component analysis (PCA) were performed revealing clear separation among three groups, with some observed overlap within the Hispanic group. Additionally, the PCA results were compared against a training set of 793 samples from the 1000 Genomes Project, demonstrating consistent results. Validation studies showed the assay to be reproducible, tolerant to common inhibitors, robust with challenging casework type samples, and sensitive down to 125 pg. In conclusion, our results demonstrated the robustness and effectiveness of a 25 loci INDEL system for ancestry inference of four ancestries commonly found in the United States.

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.

Explicature in Translating Arabic Modern Fiction into English: Protraction and Interjection

This study aims at investigating ‘explicature’ as a quadruple norm in translating Arabic modern fiction into English on the basis of the translator’s visibility. It is to show how explicature focuses on the author’s background knowledge and appreciates the reader’s language peculiarities by means of language catalysts in parentheses (i.e. interpolations). Four levels of explicature were found out: literal, formal, usable and liberal. Protracting and interjecting the potential reader’s flow of attention, the translator’s interpolations were obligatorily filling-out or specifying, optionally text-building or aesthetic, pragmatically local or global and technically inherent or revelatory. They could be modified by either full insertion, direct replacement, reverse replacement or full deletion. As the two medial types (formal and usable) are the most notable, this quadruple model of explicature is intended to be a distinct norm in translating texts of fiction across such two completely different linguistic systems and cultural backgrounds as Arabic and English.

Genetic Diversity and Population Structure Analysis of Excellent Sugar Beet (Beta vulgaris L.) Germplasm Resources

This study analyzed the genetic diversity, population structure, and cluster analysis of 129 sugar beet germplasm resources to screen superior germplasms for breeding using the 27 simple sequence repeat (SSR) and 33 pairs of insertion–deletion (InDel) molecular markers. After integrating the phenotypic variation of 16 descriptive and 4 qualitative phenotypic variables, the genetic variation levels of the 129 sugar beet germplasms’ phenotypic traits were analyzed using the principal component analysis (PCA), correlation analysis, and analysis of variance methods. The genetic diversity examination of molecular markers showed a polymorphism information content (PIC) of 0.419–0.773 (mean = 0.610). Moreover, the mean number of effective alleles detected via the SSR and InDel markers was 3.054 and 2.298, respectively. Meanwhile, the PIC ranged from 0.130 to 0.602 (mean = 0.462). The population structure analysis revealed the most appropriate K-value, indicating three populations (K = 3). The genetic distances of the 129 germplasm resources ranged from 0.099 to 0.466 (mean = 0.283). The cluster analysis results demonstrated that the germplasms were grouped into three primary classes. Based on the analysis of variance, the two qualitative features with the highest coefficients of variation were petiole width (16.64%) and length (17.11%). The descriptive trait root length index (1.395) exhibited the greatest genetic diversity. The PCA reduced the 20 phenotypic traits into five principal components, contributing 51.151%. The results of this study provide a theoretical foundation for the future selection and breeding of superior sugar beet germplasm resources.

Tracing the Maternal Line in Glacial–Interglacial Migrations of Populus tremuloides: Finding Trees for Future Sustainable Forests by Searching in the Past

Maintaining and planting sustainable forests is fundamental in perpetuating the essential functions of these ecosystems. A central aspect of managing forests for future resilience is the consideration of past migration and evolution of trees using genetic and genomic data to ensure that functionally appropriate diversity is conserved and utilized. In our study, we generated and compared genetic and genomic data from the plastome to better understand phylogeography and molecular evolution in the tree species Populus tremuloides (aspen). With these analyses, we found evidence of divergence and migration between northern and southern sites. Additionally, evidence of deep incomplete plastome sorting across the Salicaceae was found when examining insertion–deletion (indel) sites associated with DNA repair. By examining these indels in plastomic genes with introns across Salicaceae, we found a strong correlation between the abundance of DNA repair with genomic position and transcript abundance. From our findings, we conclude that previously ignored plastomic data are essential in understanding phylogeography and the evolution of key metabolic processes for improved aspen forest planning. Given the propensity of aspen forests to host high levels of biodiversity, rapidly sequester carbon, absorb excess nitrogen, and efficiently regulate snowmelt, improvements to planning and conservation will be highly impactful.

Genetic diversity analysis of big-bracted dogwood (Cornus florida and C. kousa) cultivars, interspecific hybrids, and wild-collected accessions using RADseq.

Big-bracted dogwoods are popular ornamental trees known for their beautiful spring blooms with showy bracts and four-season appeal. The two most widely grown species are Cornus florida and Cornus kousa, native to Eastern North America and East Asia. Despite their horticultural prominence, there is little information available regarding genetic diversity, population structure, relatedness, and subspecies origins of dogwood cultivars. In this study, 313 cultivars, wild-collected plants, and Rutgers University breeding selections, focusing on C. florida, C. kousa, and interspecific hybrids, were genotyped using restriction-site associated DNA sequencing (RADseq) generating thousands of single nucleotide polymorphism (SNP) and insertion deletion (Indel) markers. The research results showed high genetic diversity among C. florida and C. kousa wild-collected plants and cultivars. For C. florida, pink-bracted plants formed a distinct clade from those with white-bracts with the Mexican C. florida ssp. urbiniana forming an outgroup. For C. kousa, Chinese-collected plants (ssp. chinensis) were a distinct subspecies with clear separation from Japanese and Korean accessions (ssp. kousa) and cultivars were designated as ssp. chinensis, ssp. kousa, or ssp. hybrid. Using this information, a Kompetitive allele specific PCR (KASP) assay genotyping panel was designed to determine C. kousa trees' subspecies makeup. Results revealed many cases of genetically identical cultivars being sold under different names, especially for pink-bracted cultivars of both species. Additionally, reported parent-progeny relationships were evaluated and either validated or discredited. Finally, the hybrid germplasm analysis validated pedigrees of interspecific F1 hybrids and found many of the recent Rutgers breeding selections contain small regions of pacific dogwood (C. nuttallii) DNA introgressed into C. kousa backgrounds. This diversity study elucidates origins, diversity, and relationships of a large population of big-bracted dogwoods. The results can inform plant breeders, arboreta, and the ornamental plant industry, as most modern cultivars and popular historic cultivars are represented.

Parallel Contextual Array Insertion Deletion Grammars, Pure 2D Context-Free Grammars and Associated P Systems

Syntactic models constitute one of the main areas of mathematical studies on picture array generation. A number of 2D grammar models have been proposed with motivation from a variety of applications. Parallel Contextual Array Insertion Deletion Grammar (PCAIDG) and Pure 2D Context-Free Grammar (P2DCFG) are two such 2D grammars. In the case of P2DCFG, two new variants are studied with interesting properties. In this paper we show that PCAIDG has higher generative power than the generative powers of two variants of P2DCFG. Further, P systems associated with the grammars PCAIDG, P2DCFG and their variants are considered in the literature. In this paper, we show that the generative power of Parallel Contextual Array Insertion Deletion P System (PCAIDPS) is greater than the generative power of the P system associated with a variant of P2DCFG.

Association of insertion /deletion polymorphism of ace gene with essential hypertension in patients of Khyber Pakhtunkhwa.

The exact cause of hypertension is unknown in about 90 to 95% patients, known as essential hypertension. Genes may play a crucial role in the pathology of essential hypertension. Gene for angiotensin converting enzyme (ACE) is found on long arm of chromosome 17q23, where 287 base pair insertion or deletion (I/D) polymorphism may occur. This study was aimed to assess the association of I/D polymorphism of ACE gene with blood pressure (BP) in Patients of Khyber Pakhtunkhwa (KPK). This Descriptive Cross-sectional study was conducted from 1st June 2021 to 30th September 2021 at Kuwait Teaching Hospital, Peshawar. The genomic DNA was extracted from lymphocytes and PCR was performed for identification of ACE I/D polymorphism. Total 181 individuals (121 Hypertensive and 60 normal) were enrolled in the study. The measured systolic and diastolic BP in cases were 153.91mmHg±12.65 and 92.94mmHg±5.72, respectively while in control were 118.20±17.13 and 74.12mmHg±7.58, respectively. The Deletion Homozygous (DD), Insertion Homozygous (II) and Deletion and Insertion Heterozygous (DI) genotypes in hypertensive patients were 47 (38.84%), 17 (14.04%) and 57 (47.10%) respectively while in Control group the DD, II and DI were 4 (6.66%), 25 (41.66%) and 31 (51.66%) respectively. This study showed association of DD genotypes of the ACE gene with hypertension as compared to healthy individuals. Individuals with DD genotype may have association with hypertension. polymorphism of ACE gene was proved to be an important genetic marker for essential hypertension in Patients of KPK.

The landscape of genomic structural variation in Indigenous Australians

Indigenous Australians harbour rich and unique genomic diversity. However, Aboriginal and Torres Strait Islander ancestries are historically under-represented in genomics research and almost completely missing from reference datasets1–3. Addressing this representation gap is critical, both to advance our understanding of global human genomic diversity and as a prerequisite for ensuring equitable outcomes in genomic medicine. Here we apply population-scale whole-genome long-read sequencing4 to profile genomic structural variation across four remote Indigenous communities. We uncover an abundance of large insertion–deletion variants (20–49 bp; n = 136,797), structural variants (50 b–50 kb; n = 159,912) and regions of variable copy number (>50 kb; n = 156). The majority of variants are composed of tandem repeat or interspersed mobile element sequences (up to 90%) and have not been previously annotated (up to 62%). A large fraction of structural variants appear to be exclusive to Indigenous Australians (12% lower-bound estimate) and most of these are found in only a single community, underscoring the need for broad and deep sampling to achieve a comprehensive catalogue of genomic structural variation across the Australian continent. Finally, we explore short tandem repeats throughout the genome to characterize allelic diversity at 50 known disease loci5, uncover hundreds of novel repeat expansion sites within protein-coding genes, and identify unique patterns of diversity and constraint among short tandem repeat sequences. Our study sheds new light on the dimensions and dynamics of genomic structural variation within and beyond Australia.

Complete genomic profiles of 1,496 Taiwanese reveal curated medical insights

IntroductionThe population of Taiwan has a long history of ethno-cultural evolution. The Taiwanese population was isolated from other large populations such as the European, Han Chinese, and Japanese population. The Taiwan Biobank (TWB) project has built a nationwide database, particularly for personal whole-genome sequence (WGS) to facilitate basic and clinical collaboration nationally and internationally, making it one of the most valuable public datasets of the East Asian population. ObjectivesThis study provides comprehensive medical genomic findings from TWB WGS data, for better characterization of disease susceptibility and the choice of ideal treatment regimens in Taiwanese population. MethodsWe reanalyzed 1,496 WGS using a PrecisionFDA Truth challenge winner method Sentieon DNAscope. Single nucleotide variants (SNV) and small insertions/deletions (INDEL) were benchmarked. We also analyzed pharmacogenomic (PGx) drug-associated alleles, and copy number variants (CNV). Multiple practicing clinicians reviewed and curated the clinically significant variants. Variant annotations can be browsed at TaiwanGenomes (https://genomes.tw). ResultsWe found that each participant had an average of 6,870.7 globally novel variants and 75.3% (831/1103) of the participants harbored at least one PharmGKB-selected high evidence level human leukocyte antigen (HLA) risk allele. 54 PharmGKB-reported high-level instances of evidence of Cytochrome P450 variant-drug pairs, with a population frequency of over 13.2%. We also identified 23 variants in the ACMG secondary finding V3 gene list from 25 participants, suggesting that 1.67% (25/1496) of the population is harboring at least one medical actionable variant. Our carrier status analyses suggest that one in 25 couples (3.94%) would risk having offspring with at least one pathogenic variant, which is in line with rates found in Japan and Singapore. For pathogenic CNV, we detected 6.88% and 2.02% carrier rates for alpha thalassemia and spinal muscular atrophy, respectively. ConclusionOur study highlights the overall medical insights of a complete Taiwanese genomic profile.

Transposable elements cause the loss of self-incompatibility in citrus.

Self-incompatibility (SI) is a widespread prezygotic mechanism for flowering plants to avoid inbreeding depression and promote genetic diversity. Citrus has an S-RNase-based SI system, which was frequently lost during evolution. We previously identified a single nucleotide mutation in Sm-RNase, which is responsible for the loss of SI in mandarin and its hybrids. However, little is known about other mechanisms responsible for conversion of SI to self-compatibility (SC) and we identify a completely different mechanism widely utilized by citrus. Here, we found a 786-bp miniature inverted-repeat transposable element (MITE) insertion in the promoter region of the FhiS2-RNase in Fortunella hindsii Swingle (a model plant for citrus gene function), which does not contain the Sm-RNase allele but are still SC. We demonstrate that this MITE plays a pivotal role in the loss of SI in citrus, providing evidence that this MITE insertion prevents expression of the S-RNase; moreover, transgenic experiments show that deletion of this 786-bp MITE insertion recovers the expression of FhiS2-RNase and restores SI. This study identifies the first evidence for a role for MITEs at the S-locus affecting the SI phenotype. A family-wide survey of the S-locus revealed that MITE insertions occur frequently adjacent to S-RNase alleles in different citrus genera, but only certain MITEs appear to be responsible for the loss of SI. Our study provides evidence that insertion of MITEs into a promoter region can alter a breeding strategy and suggests that this phenomenon may be broadly responsible for SC in species with the S-RNase system.

2160. Emergence of INDEL Mutations in KPC-2: Insights into Ceftazidime-Avibactam and Imipenem-Relebactam Efficacy

Abstract Background Enterobacterales producing KPC-2 and KPC-3 β-lactamases (Bla) are a significant clinical threat. Ceftazidime/avibactam (CZA) and imipenem/relebactam (IMI/REL) are successful in the treatment for these infections. Unfortunately, CZA-resistant isolates are emerging as a result of amino acid changes in three “hotspots” in the KPC Bla: Ω-loop region; the 237-243 loop; and the 266-275 loop (Figure 1). Our goal here is to assess the effect of insertion-deletion (INDEL) mutations located in the three hotspots in KPC-2, and their impact on the efficacy of these two drug combinations Methods The blaKPC-44 (ins_261_AVYTRAPNKDDKHSE), blaKPC-80 (ins_267_PNK), blaKPC-81 (del_173_I), blaKPC-97 (ins_276_VNSEA), and blaKPC-160 (del_167_LE) genes were amplified by PCR from K. pneumoniae isolates resistant to CZA. The blaKPC-14 (del_242-243_GT) gene was obtained by mutagenesis from the blaKPC-2 gene. Genes were cloned in pMBLe and transformed in E. coli TOP10 for MIC determinations. Immunoblotting with polyclonal anti-KPC-2 antibody was performed in whole-cell preparations. Molecular modeling of KPC variants were obtained by Yasara Results All KPC variants demonstrated lower MICs compared to KPC-2 for most β-lactams (carbapenems included), except for CAZ, where variants exhibited up to 8-fold higher MICs. All variant clones showed at least a 3-dilution higher MICs for CZA, except KPC-160. IMI-REL restored susceptibility to KPC-2 and INDEL variants. The Ω−loop variants exhibited lower MICs compared to the other hotspots variants (CZA included). INDEL mutations resulted in decreased levels of protein production, with KPC-81 production being the most affected. In silico models suggest that INDEL mutations have different effects depending on the structural domain affectedTable 1.Minimum Inhibitory Concentrations (μg/mL) of E. coli TOP10F´ recombinant clones Conclusion INDEL mutations in the three hotspots of KPC-2 can impair CZA activity, but not affect IMI/REL susceptibility. The differences in MICs among the variants suggest that each hotspot affects the interactions of KPC-2 with CAZ and AVI through different mechanisms. Additionally, INDEL mutations appear to decrease protein stability as Bla production levels are reduced. This work provides relevant information about the changes in the interaction with DBO inhibitors associated with INDEL mutations in KPC-2 Disclosures Robert A. bonomo, MD, Entasis, Merck, VenatoRx, Wockhardt: Grant/Research Support

Pneumonic Plague Protection Induced by a Monophosphoryl Lipid A Decorated Yersinia Outer-Membrane-Vesicle Vaccine.

A new Yersinia pseudotuberculosis mutant strain, YptbS46, carrying the lpxE insertion and pmrF-J deletion is constructed and shown to exclusively produce monophosphoryl lipid A (MPLA) having adjuvant properties. Outer membrane vesicles (OMVs) isolated from YptbS46 harboring an lcrV expression plasmid, pSMV13, are designated OMV46-LcrV, which contained MPLA and high amounts of LcrV (Low Calcium response V) and displayed low activation of Toll-like receptor 4 (TLR4). Intramuscular prime-boost immunization with 30µg of of OMV46-LcrV exhibited substantially reduced reactogenicity than the parent OMV44-LcrV and conferred complete protection to mice against a high-dose of respiratory Y. pestis challenge. OMV46-LcrV immunization induced robust adaptive responses in both lung mucosal and systemic compartments and orchestrated innate immunity in the lung, which are correlated with rapid bacterial clearance and unremarkable lung damage during Y. pestis challenge. Additionally, OMV46-LcrV immunization conferred long-term protection. Moreover, immunization with reduced doses of OMV46-LcrV exhibited further lower reactogenicity and still provided great protection against pneumonic plague. The studies strongly demonstrate the feasibility of OMV46-LcrV as a new type of plague vaccine candidate.