Single Gene Locus Research Articles

Enzyme Machinery for Bacterial Glucoside Metabolism through a Conserved Non-hydrolytic Pathway.

The flexible acquisition of substrates from nutrient pools is critical for microbes to prevail in competitive environments. To acquire glucose from diverse glycoside and disaccharide substrates, many free-living and symbiotic bacteria have developed, alongside hydrolysis, a non-hydrolytic pathway comprised of four biochemical steps and conferred from a single glycoside utilization gene locus (GUL). Mechanistically, this pathway integrates within the framework of oxidation and reduction at the glucosyl/glucose C3, the eliminative cleavage of the glycosidic bond and the addition of water in two consecutive lyase-catalyzed reactions. Here, based on study of enzymes from the phytopathogen Agrobacterium tumefaciens, we reveal a conserved Mn2+ metallocenter active site in both lyases and identify the structural requirements for specific catalysis to elimination of 3-keto-glucosides and water addition to the resulting 2-hydroxy-3-keto-glycal product, yielding 3-keto-glucose. Extending our search of GUL-encoded putative lyases to the human gut commensal Bacteroides thetaiotaomicron, we discover a Ca2+ metallocenter active site in a putative glycoside hydrolase-like protein and demonstrate its catalytic function in the eliminative cleavage of 3-keto-glucosides of opposite (α) anomeric configuration as preferred by the A. tumefaciens enzyme (β). Structural and biochemical comparisons reveal the molecular-mechanistic origin of 3-keto-glucoside lyase stereo-complementarity. Our findings identify a basic set of GUL-encoded lyases for glucoside metabolism and assign physiological significance to GUL genetic diversity in the bacterial domain of life.

Enzymmaschinerie für bakteriellen Glucosidmetabolismus über einen konservierten nicht‐hydrolytischen Abbauweg

AbstractFlexible Substrataufnahme aus Nährstoffreservoirs ist für Mikroorganismen entscheidend, um sich in kompetitiven natürlichen Umgebungen durchzusetzen. Zur Aufnahme von Glucose aus verschiedenen Glucosid‐ und Disaccharid‐Substraten haben viele frei‐ und symbiotisch lebende Bakterien, neben der Hydrolyse, einen nicht‐hydrolytischen Stoffwechselweg entwickelt. Dieser besteht aus vier biochemischen Schritten und wird von einem distinkten Genlocus (glycoside utilization locus, GUL) codiert. Mechanistisch integriert der Stoffwechselweg die Oxidation und Reduktion am Glucosyl/Glucose‐C3 mit der eliminativen Spaltung der glycosidischen Bindung und der Addition von Wasser an das Eliminationsprodukt in zwei aufeinanderfolgenden, von Lyasen katalysierten Reaktionen. In dieser Studie zeigen wir, basierend auf der Untersuchung von Enzymen des Pflanzenpathogens Agrobacterium tumefaciens, ein konserviertes Aktivitätszentrum in beiden Lyasen, welches um ein zentrales Mn2+ Metallion angeordnet ist. Weiters identifizieren wir die strukturellen Voraussetzungen für die spezifische Katalyse der 3‐Ketoglucosid Eliminierung zu 2‐Hydroxy‐3‐ketoglycal und der Hydratisierung des Glycal zu 3‐Ketoglucose. Die Erweiterung unserer Suche nach GUL‐codierten putativen Lyasen auf das menschliche darmkommensale Bakterium Bacteroides thetaiotaomicron führte uns zur Entdeckung eines Ca2+‐koordinierten Aktivitätszentrums in einem Glycosidase‐ähnlichen Protein. Das Enzym katalysiert die eliminative Spaltung von 3‐Ketoglucosiden entgegengesetzter (α) anomerer Konfiguration im Vergleich zu den von der A. tumefaciens Lyase präferierten Substraten (β). Strukturelle und biochemische Vergleiche zeigen die molekular‐mechanistische Basis der Stereokomplementarität der beiden 3‐Ketoglucosid Lyasen. Unsere Ergebnisse identifizieren ein grundlegendes Set von GUL‐codierten Lyasen für den Glucosidmetabolismus und erlauben es, der Gendiversität von GULs in der Domäne von Bakterien physiologische Bedeutung zuzuschreiben.

Analysis of Fluorescent Proteins for Observing Single Gene Locus in a Live and Fixed Escherichia coli Cell.

Fluorescent proteins (FPs) are essential tools for advanced microscopy techniques such as super-resolution imaging, single-particle tracking, and quantitative single-molecule counting. Various FPs fused to DNA-binding proteins have been used to observe the subcellular location and movement of specific gene loci in living and fixed bacterial cells. However, quantitative assessments of the properties of FPs for gene locus measurements are still lacking. Here, we assessed various FPs to observe specific gene loci in live and fixed Escherichia coli cells using a fluorescent repressor-operator binding system (FROS), tet operator-Tet repressor proteins (TetR). Tsr-fused FPs were used to assess the intensity and photostability of various FPs (five red FPs: mCherry2, FusionRed, mRFP, mCrimson3, and dKatushka; and seven yellow FPs: SYFP2, Venus, mCitrine, YPet, mClover3, mTopaz, and EYFP) at the single-molecule level in living cells. These FPs were then used for gene locus measurements using FROS. Our results indicate that TetR-mCrimson3 (red) and TetR-EYFP (yellow) had better properties for visualizing gene loci than the other TetR-FPs. Furthermore, fixation procedures affected the clustering of diffusing TetR-FPs and altered the locations of the TetR-FP foci. Fixation with formaldehyde consistently disrupted proper DNA locus observations using TetR-FPs. Notably, the foci measured using TetR-mCrimson3 remained close to their original positions in live cells after glyoxal fixation. This in vivo study provides a cell-imaging guide for the use of FPs for gene-locus observation in E. coli and a scheme for evaluating the use of FPs for other cell-imaging purposes.

Correlation analysis between driver gene mutation and clinicopathological features in lung adenocarcinoma based on real-world cumulative clinical data.

Driver genes are essential predictors of targeted therapeutic efficacy. Detecting driver gene mutations in lung adenocarcinoma (LUAD) patients can help to screen for targeted drugs and improve patient survival benefits. This study aims to investigate the mutation characterization of driver genes and their correlation with clinicopathological features in LUAD. A total of 440 LUAD patients were selected from Sir Run Run Shaw Hospital between July 2019 and September 2022. Postoperative tissue specimens were analyzed for gene mutations using next-generation sequencing technology, focusing, including epidermal growth factor receptor EGFR, ALK, ROS1, RET, KRAS, MET, BRAF, HER2, PIK3CA and NRAS. At the same time, clinicopathological data were collected and organized for multidimensional correlation analysis. Of 440 LUAD patients, driver gene mutations were not detected in 48 patients. The proportion of patients with driver gene mutations was as high as 89.09%. The top three driver genetic mutations were EGFR, KRAS, and MET. Sixty-nine types of EGFR mutations were detected and distributed in the protein tyrosine kinase catalytic domain (56, 81.16%), Furin-like cysteine-rich region (9, 13.04%), receptor binding domain (3, 4.35%), and EGFR transmembrane domain (1, 1.45%). Single gene locus mutation occurred in 343 LUAD patients, but the mutation gene types covered all tested genes. Our findings showed that EGFR mutations were more commonly observed in non-smoking and female patients (P<0.01), KRAS mutations were more prevalent in male patients and smokers (P<0.01), ROS1 mutations had larger tumor diameters (P<0.01) and RET mutations were more prevalent in smokers (P<0.05). LUAD patients exhibit diverse genetic mutations, which may co-occur simultaneously. Integrated analysis of multiple mutations is essential for accurate diagnosis and effective treatment of the disease. The use of NGS can significantly expand our understanding of gene mutations and facilitate integrated analysis of multiple gene mutations, providing critical evidence for targeted treatment methods.

A gene with a thousand alleles: The hyper-variable effectors of plant-parasitic nematodes

Pathogens are engaged in a fierce evolutionary arms race with their host. The genes at the forefront of the engagement between kingdoms are often part of diverse and highly mutable gene families. Even in this context, we discovered unprecedented variation in the hyper-variable (HYP) effectors of plant-parasitic nematodes. HYP effectors are single-gene loci that potentially harbor thousands of alleles. Alleles vary in the organization, as well as the number, of motifs within a central hyper-variable domain (HVD). We dramatically expand the HYP repertoire of two plant-parasitic nematodes and define distinct species-specific "rules" underlying the apparently flawless genetic rearrangements. Finally, by analyzing the HYPs in 68 individual nematodes, we unexpectedly found that despite the huge number of alleles, most individuals are germline homozygous. These data support a mechanism of programmed genetic variation, termed HVD editing, where alterations are locus specific, strictly governed by rules, and theoretically produce thousands of variants without errors.

#3001 MtDNA copy number correlates to coronary artery calcium score and biological age in kidney failure patients

Abstract Background and Aims Chronic kidney disease (CKD) and kidney failure are connected to increased oxidative stress and vascular calcification, which are indicators for development of comorbidities and mortality outcome. Mitochondrial (mt)DNA copy number has been reported as independent predictor for frailty and mortality outcome of various cardiovascular diseases (CVD) and cancer. We aimed that mtDNA copy number in whole blood of kidney failure patients can be correlated to coronary artery media calcification score and coronary artery calcium (CAC) score and be used as an independent predictor for comorbidities and mortality outcome. Method Quantitative PCR (qPCR) with TaqMan® probes for three mtDNA genes (mtND1, mtND4 and mtCOX1) and two single locus genes on nuclear DNA (hemoglobin subunit beta and 18S), for normalization, is performed on DNA samples isolated from collected blood of 196 kidney transplant donors and 211 recipients at basal timepoint and 32 at 1-year-follow-up timepoint, included in the KaroKidney biobank. Resulting mtDNA copy number is investigated for correlation to CAC score, patient mortality outcome, biological age determined by skin autofluorescence and other clinical parameters. Results In our Swedish kidney transplant cohort we can see statistically significant difference in mtDNA copy number between kidney failure patients (recipients) and healthy controls (donors). No obvious difference between females and males was observed. mtDNA copy number correlates to biological age determined by skin autofluorescence, CAC score and Framingham CVD risk score. In the 32 1-year-follow-up samples we could see significant increase/recovery of mtDNA copy number for male patients compared to their baseline before receiving a transplant. Conclusion In this study we show that mtDNA copy number in kidney failure patients correlates to biological age and CAC score, affirming mtDNA copy number as an independent predictor of frailty in this Swedish cohort of CKD patients. Additionally, we can see that a kidney transplant can lead to a significant increase of mtDNA copy number in blood one year later, suggesting a general increase in health quality.

Genome structural dynamics: insights from Gaussian network analysis of Hi-C data.

Characterization of the spatiotemporal properties of the chromatin is essential to gaining insights into the physical bases of gene co-expression, transcriptional regulation and epigenetic modifications. The Gaussian network model (GNM) has proven in recent work to serve as a useful tool for modeling chromatin structural dynamics, using as input high-throughput chromosome conformation capture data. We focus here on the exploration of the collective dynamics of chromosomal structures at hierarchical levels of resolution, from single gene loci to topologically associating domains or entire chromosomes. The GNM permits us to identify long-range interactions between gene loci, shedding light on the role of cross-correlations between distal regions of the chromosomes in regulating gene expression. Notably, GNM analysis performed across diverse cell lines highlights the conservation of the global/cooperative movements of the chromatin across different types of cells. Variations driven by localized couplings between genomic loci, on the other hand, underlie cell differentiation, underscoring the significance of the four-dimensional properties of the genome in defining cellular identity. Finally, we demonstrate the close relation between the cell type-dependent mobility profiles of gene loci and their gene expression patterns, providing a clear demonstration of the role of chromosomal 4D features in defining cell-specific differential expression of genes.

Alternative polyadenylation regulates acetyl-CoA carboxylase function in peanut

BackgroundPolyadenylation is a crucial process that terminates mRNA molecules at their 3′-ends. It has been observed that alternative polyadenylation (APA) can generate multiple transcripts from a single gene locus, each with different polyadenylation sites (PASs). This leads to the formation of several 3′ untranslated regions (UTRs) that vary in length and composition. APA has a significant impact on approximately 60–70% of eukaryotic genes and has far-reaching implications for cell proliferation, differentiation, and tumorigenesis.ResultsIn this study, we conducted long-read, single-molecule sequencing of mRNA from peanut seeds. Our findings revealed that over half of all peanut genes possess over two PASs, with older developing seeds containing more PASs. This suggesting that the PAS exhibits high tissue specificity and plays a crucial role in peanut seed maturation. For the peanut acetyl-CoA carboxylase A1 (AhACCA1) gene, we discovered four 3′ UTRs referred to UTR1–4. RT-PCR analysis showed that UTR1-containing transcripts are predominantly expressed in roots, leaves, and early developing seeds. Transcripts containing UTR2/3 accumulated mainly in roots, flowers, and seeds, while those carrying UTR4 were constitutively expressed. In Nicotiana benthamiana leaves, we transiently expressed all four UTRs, revealing that each UTR impacted protein abundance but not subcellular location. For functional validation, we introduced each UTR into yeast cells and found UTR2 enhanced AhACCA1 expression compared to a yeast transcription terminator, whereas UTR3 did not. Furthermore, we determined ACC gene structures in seven plant species and identified 51 PASs for 15 ACC genes across four plant species, confirming that APA of the ACC gene family is universal phenomenon in plants.ConclusionOur data demonstrate that APA is widespread in peanut seeds and plays vital roles in peanut seed maturation. We have identified four 3′ UTRs for AhACCA1 gene, each showing distinct tissue-specific expression patterns. Through subcellular location experiment and yeast transformation test, we have determined that UTR2 has a stronger impact on gene expression regulation compared to the other three UTRs.

New evidence of consistency between phylogeny and morphology for two taxa in ciliated protists, the subclasses Oligotrichia and Choreotrichia (Protista, Ciliophora)

Marine planktonic ciliates are largely oligotrichs and choreotrichs, which are two subclasses of the class Spirotrichea. The current phylogenetic assignments of oligotrichs and choreotrichs are inconsistent with previous results based on morphological features, probably hindered by the limited information from a single gene locus. Here we provide 53 new sequences from small subunit ribosomal RNA (SSU rDNA), ITS1-5.8S rDNA-ITS2, and large subunit ribosomal RNA (LSU rDNA) gene loci in 25 oligotrich and choreotrich species. We also predict RNA secondary structures for the ITS2 regions in 55 species, 48 species of which are reported for the first time. Based on these novel data, we make a more comprehensive phylogenetic reconstruction, revealing consistency between morphological taxonomy and an updated phylogenetic system for oligotrichs and choreotrichs. With the addition of data from ciliature patterns and genes, the phylogenetic analysis of the subclass Oligotrichia suggests three evolutionary trajectories, among which: 1) Novistrombidium asserts an ancestral ciliary pattern in Oligotrichia; 2) the subgenera division of Novistrombidium and Parallelostrombidium are fully supported; 3) the three families (Tontoniidae, Pelagostrombidiidae and Cyrtostrombidiidae) all evolved from the most diverse family Strombidiidae, which explains why strombidiids consistently form polyphyletic clades. In the subclass Choreotrichia, Strombidinopsis likely possesses an ancestral position to other choreotrichs, and both phylogenetic analysis and RNA secondary structure prediction support the hypothesis that tintinnids may have evolved from Strombidinopsis. The results presented here offer an updated hypothesis for the evolutionary history of oligotrichs and choreotrichs based on new evidence obtained by expanding sampling of molecular information across multiple gene loci.

Comparative Transcriptome Analysis Reveals Novel Candidate Resistance Genes Involved in Defence against Phytophthora cactorum in Strawberry.

Crown rot, caused by Phytophthora cactorum, is a devastating disease of strawberry. While most commercial octoploid strawberry cultivars (Fragaria × ananassa Duch) are generally susceptible, the diploid species Fragaria vesca is a potential source of resistance genes to P. cactorum. We previously reported several F. vesca genotypes with varying degrees of resistance to P. cactorum. To gain insights into the strawberry defence mechanisms, comparative transcriptome profiles of two resistant genotypes (NCGR1603 and Bukammen) and a susceptible genotype (NCGR1218) of F. vesca were analysed by RNA-Seq after wounding and subsequent inoculation with P. cactorum. Differential gene expression analysis identified several defence-related genes that are highly expressed in the resistant genotypes relative to the susceptible genotype in response to P. cactorum after wounding. These included putative disease resistance (R) genes encoding receptor-like proteins, receptor-like kinases, nucleotide-binding sites, leucine-rich repeat proteins, RPW8-type disease resistance proteins, and 'pathogenesis-related protein 1'. Seven of these R-genes were expressed only in the resistant genotypes and not in the susceptible genotype, and these appeared to be present only in the genomes of the resistant genotypes, as confirmed by PCR analysis. We previously reported a single major gene locus RPc-1 (Resistance to Phytophthora cactorum 1) in F. vesca that contributed resistance to P. cactorum. Here, we report that 4-5% of the genes (35-38 of ca 800 genes) in the RPc-1 locus are differentially expressed in the resistant genotypes compared to the susceptible genotype after inoculation with P. cactorum. In particular, we identified three defence-related genes encoding wall-associated receptor-like kinase 3, receptor-like protein 12, and non-specific lipid-transfer protein 1-like that were highly expressed in the resistant genotypes compared to the susceptible one. The present study reports several novel candidate disease resistance genes that warrant further investigation for their role in plant defence against P. cactorum.

A genetic Study to Darwin's tubercle trait of External ear in the Population of Salah Al –Dein and Kirkuk

A genetic study for the trait of Darwin's tubercle in two ways : first , on a sample consisted of 52 family from Tikrit city in governorate Salah AL-Dein and 125 individual from their sons and 63 family from governorate of Kirkuk and 188 individual from their sons . Secondly , adoption of pedigree analysis method for seven families from governorate of Salah El-Dein and Kirkuk , which confirmed the darwin's tubercle trait is a genetic trait and the presence of tubercle is dominant absence . The results of Presence Darwin's tubercle also revealed different forms : Nodosity tubercle , Enlargement tubercle and Projection. The present study support the truth of single gene locus hypothesis for the inheritance of this trait , in spite of it record the presence of one offspring have darwin's tubercle between two parents do not have darwin's tubercle in one family. The results of the study distribution of phenotypic for the trait showed on a sample consisted of 1415 individual from the citizens of the governorate of Salah Al-Dein (920♂: 495♀) from the original samples (1446) , an elevated percentage of the presence tubercle among the males (99.2%) than the females (98.4%) . The percentage of presence tubercle in the whole population was (98.9%) . Concering the allelic frequency for the gene of darwin's tubercle, the D allele was observed a higher frequency than the d allele in the whole population of the governorate: 0.895, 0.105 for both of them respectively . Expected frequencies of the three genotypes of darwin's tubercle trait , revealed a prevalence of the homozygous (dominant) genotype DD over the homozygous (recessive) and heterozygous genotype in the population of the governorate: 0.801, 0.188, 0.011 for DD , Dd and dd respectively.

Imaging-Based In Situ Analysis of 5-Methylcytosine at Low Repetitive Single Gene Loci with Transcription-Activator-Like Effector Probes.

Transcription-activator-like effectors (TALEs) are programmable DNA binding proteins that can be used for sequence-specific, imaging-based analysis of cellular 5-methylcytosine. However, this has so far been limited to highly repetitive satellite DNA. To expand this approach to the analysis of coding single gene loci, we here explore a number of signal amplification strategies for increasing imaging sensitivity with TALEs. We develop a straightforward amplification protocol and employ it to target the MUC4 gene, which features only a small cluster of repeat sequences. This offers high sensitivity imaging of MUC4, and in costaining experiments with pairs of one TALE selective for unmethylated cytosine and one universal control TALE enables analyzing methylation changes in the target independently of changes in target accessibility. These advancements offer prospects for 5-methylcytosine analysis at coding, nonrepetitive gene loci by the use of designed TALE probe collections.

Implications of differential transcription start site selection on chronic myeloid leukemia and prostate cancer cell protein expression.

The relevance of minor transcription start sites in broad promoters is not well understood. We have studied AGAP2 expression in prostate cancer and chronic myeloid leukemia (CML), showing transcription is initiated from alternative transcription start sites (TSSs) within a single TSS cluster, producing cancer-type-specific AGAP2 mRNAs with small differences in their 5' UTR length. Interestingly, in the CML cell lines where the 5' UTR is longer, AGAP2 protein levels are lower. We demonstrate that the selection of an upstream TSS involved the formation of a G quadruplex in the 5' UTR, decreasing polysome formation. After developing a bioinformatics pipeline to query data from the FANTOM project and the NCl-60 human tumor cell lines screen, we found HK1 expression can also be regulated by the same mechanism. Overall, we present compelling data supporting TSS selection within a TSS cluster play a role in protein expression and should not be ignored.

Plant speciation in the Namib Desert: potential origin of a widespread derivative species from a narrow endemic

ABSTRACT Background Parapatric (or ‘budding’) speciation is increasingly recognised as an important phenomenon in plant evolution but its role in extreme (e.g. desert) environments is poorly documented. Aims To test this speciation model in a hypothesised sister pair, the Southwest – North African disjunct Senecio flavus and its putative progenitor, the Namibian Desert endemic S. englerianus. Methods Phylogenetic inferences were combined with niche divergence tests, morphometrics, and experimental-genetic approaches. We also evaluated the potential role of an African Dry Corridor (ADC) in promoting the hypothesised northward expansion of S. flavus (from Namibia), using palaeodistribution models. Results Belonging to an isolated (potential ‘relict’) clade, the two morphologically distinct species showed pronounced niche divergence in Namibia and signs of digenic-epistatic hybrid incompatibility (based on F2 pollen fertility). The presence of ‘connate-fluked’ pappus hairs in S. flavus, likely increasing dispersal ability, is controlled by a single-gene locus. Conclusions Our results provide evidence for a possible (and rare) example of ‘budding’ speciation in which a wider-ranged derivative (S. flavus) originated at the periphery of a smaller-ranged progenitor (S. englerianus) in the Namib Desert region. The Southwest – North African disjunction of S. flavus could have been established by dispersal across intermediate ADC areas during periods of (Late) Pleistocene aridification.

Familial posterior predominant subcortical band heterotopia caused by a CEP85L missense mutation

Lissencephaly comprises a spectrum of cortical malformations caused by disruption of neuronal migration. The lissencephaly spectrum includes agyria defined as cortex lacking gyri with sulci >3 cm apart, pachygyria defined as abnormally broad gyri with sulci 1.5–3 cm apart, and subcortical band heterotopia (SBH) characterised by a band of heterotopic neurons beneath the cortex, separated by a thin layer of white matter [1]. Lissencephaly has heterogeneous genetic aetiologies, including both single gene mutations (for example, LIS1 and DCX variants) and locus deletions (for example, 17p13.3 deletion causing Miller-Dieker syndrome) [1].

Microevolution of Trypanosoma cruzi reveals hybridization and clonal mechanisms driving rapid genome diversification.

Protozoa and fungi are known to have extraordinarily diverse mechanisms of genetic exchange. However, the presence and epidemiological relevance of genetic exchange in Trypanosoma cruzi, the agent of Chagas disease, has been controversial and debated for many years. Field studies have identified both predominantly clonal and sexually recombining natural populations. Two of six natural T. cruzi lineages (TcV and TcVI) show hybrid mosaicism, using analysis of single-gene locus markers. The formation of hybrid strains in vitro has been achieved and this provides a framework to study the mechanisms and adaptive significance of genetic exchange. Using whole genome sequencing of a set of experimental hybrids strains, we have confirmed that hybrid formation initially results in tetraploid parasites. The hybrid progeny showed novel mutations that were not attributable to either (diploid) parent showing an increase in amino acid changes. In long-term culture, up to 800 generations, there was a variable but gradual erosion of progeny genomes towards triploidy, yet retention of elevated copy number was observed at several core housekeeping loci. Our findings indicate hybrid formation by fusion of diploid T. cruzi, followed by sporadic genome erosion, but with substantial potential for adaptive evolution, as has been described as a genetic feature of other organisms, such as some fungi.

NAD+ deficiency in human congenital malformations and miscarriage: A new model of pleiotropy.

Pleiotropy is defined as the phenomenon of a single gene locus influencing two or more distinct phenotypic traits. However, nicotinamide adenine dinucleotide (NAD+) deficiency through diet alone can cause multiple or single malformations in mice. Additionally, humans with decreased NAD+ production due to changes in pathway genes display similar malformations. Here, I hypothesize NAD+ deficiency as a pleiotropic mechanism for multiple malformation conditions, including limb‐body wall complex (LBWC), pentalogy of Cantrell (POC), omphalocele‐exstrophy‐imperforate anus‐spinal defects (OEIS) complex, vertebral‐anal‐cardiac‐tracheoesophageal fistula‐renal‐limb (VACTERL) association (hereafter VACTERL), oculoauriculovertebral spectrum (OAVS), Mullerian duct aplasia‐renal anomalies‐cervicothoracic somite dysplasia (MURCS), sirenomelia, and urorectal septum malformation (URSM) sequence, along with miscarriages and other forms of congenital malformation. The term Congenital NAD Deficiency Disorder (CNDD) could be considered for patients with these malformations; however, it is important to emphasize there have been no confirmatory experimental studies in humans to prove this hypothesis. In addition, these multiple malformation conditions should not be considered individual entities for the following reasons: First, there is no uniform consensus of clinical diagnostic criteria and all of them fail to capture cases with partial expression of the phenotype. Second, reports of individuals consistently show overlapping features with other reported conditions in this group. Finally, what is currently defined as VACTERL is what I would refer to as a default label when more striking features such as body wall defects, caudal dysgenesis, or cloacal exstrophy are not present.

Simulation Research on the Methods of Multi-Gene Region Association Analysis Based on a Functional Linear Model.

Genome-wide association analysis is an important approach to identify genetic variants associated with complex traits. Complex traits are not only affected by single gene loci, but also by the interaction of multiple gene loci. Studies of association between gene regions and quantitative traits are of great significance in revealing the genetic mechanism of biological development. There have been a lot of studies on single-gene region association analysis, but the application of functional linear models in multi-gene region association analysis is still less. In this paper, a functional multi-gene region association analysis test method is proposed based on the functional linear model. From the three directions of common multi-gene region method, multi-gene region weighted method and multi-gene region loci weighted method, that test method is studied combined with computer simulation. The following conclusions are obtained through computer simulation: (a) The functional multi-gene region association analysis test method has higher power than the functional single gene region association analysis test method; (b) The functional multi-gene region weighted method performs better than the common functional multi-gene region method; (c) the functional multi-gene region loci weighted method is the best method for association analysis on three directions of the common multi-gene region method; (d) the performance of the Step method and Multi-gene region loci weighted Step for multi-gene regions is the best in general. Functional multi-gene region association analysis test method can theoretically provide a feasible method for the study of complex traits affected by multiple genes.

Kastor and Polluks polypeptides encoded by a single gene locus cooperatively regulate VDAC and spermatogenesis

Although several long noncoding RNAs (lncRNAs) have recently been shown to encode small polypeptides, those in testis remain largely uncharacterized. Here we identify two sperm-specific polypeptides, Kastor and Polluks, encoded by a single mouse locus (Gm9999) previously annotated as encoding a lncRNA. Both Kastor and Polluks are inserted in the outer mitochondrial membrane and directly interact with voltage-dependent anion channel (VDAC), despite their different amino acid sequences. Male VDAC3-deficient mice are infertile as a result of reduced sperm motility due to an abnormal mitochondrial sheath in spermatozoa, and deficiency of both Kastor and Polluks also severely impaired male fertility in association with formation of a similarly abnormal mitochondrial sheath. Spermatozoa lacking either Kastor or Polluks partially recapitulate the phenotype of those lacking both. Cooperative function of Kastor and Polluks in regulation of VDAC3 may thus be essential for mitochondrial sheath formation in spermatozoa and for male fertility.

Application of Next-Generation Sequencing Technology Based on Single Gene Locus in Species Identification of Mixed Meat Products

Polymerase chain reaction (PCR) detection is a commonly used method for species identification of meat products. However, this method is not suitable for the analysis of meat products containing multiple mixtures. This study aimed to test whether next-generation sequencing (NGS) technology could be used as a method for the certification of mixed meat products. In this study, five kinds of common meat (pigs, cattle, sheep, chickens, and ducks) were mixed as samples with different proportions. The primers designed from mitochondrial 16S rRNA and nuclear genome gene (growth hormone receptor, GHR), respectively, were used to detect these meats. The sequencing results of NGS were analyzed using a self-designed bioinformatics program. The fragments with similar sequences were classified and compared with the database to determine their species. The results showed that all five kinds of meat components could be correctly identified using these two primers. The meat composition could be detected as low as 0.5% in the mixed samples using the NGS technology targeting GHR gene fragments, which was superior to those targeting mitochondrial 16S rRNA. However, the quantitative detection of species in the mixture was not likely to be quite accurate due to the amplification bias of PCR amplification. These results showed that the NGS technology could be applied to identify meat species in mixtures.