Marker-assisted Selection Research Articles

Exploring crop genomics: role of molecular markers in identifying genetic diversity and characterization

and facilitating germplasm characterization for crop improvement. Understanding and harnessing crop genetic diversity are paramount with the escalating demand for sustainable agricultural practices and the need to address global food security challenges. Molecular markers have emerged as powerful tools enabling precise identification, characterization, and utilization of genetic resources for crop enhancement. The paper provides an in-depth analysis of various molecular marker techniques, encompassing DNA-based markers such as single nucleotide polymorphisms (SNPs), simple sequence repeats (SSRs), and amplified fragment length polymorphisms (AFLPs), as well as RNA-based markers like expression sequence tags (ESTs) and microRNAs. By elucidating genetic relationships, population structure, and phylogenetic analysis, these markers facilitate the conservation and utilization of crop germplasm. Moreover, the review highlights recent advancements in high-throughput genotyping technologies and bioinformatics tools, which have revolutionized crop genomic research. These advancements enable comprehensive genome-wide analyses, accelerating breeding efforts for the development of improved crop varieties with enhanced traits such as yield, quality, and stress tolerance. Integration of molecular marker-assisted selection (MAS) into breeding programs further enhances the efficiency and precision of crop improvement strategies. Despite the significant contributions of molecular markers to crop genomics, challenges persist. The review addresses these challenges and discusses potential solutions, emphasizing the importance of collaborative efforts, data sharing, and interdisciplinary research endeavors. By overcoming these hurdles, the agricultural community can fully harness the potential of genomics for sustainable crop improvement, thus addressing global food security challenges in the face of changing environmental conditions and population growth. In conclusion, understanding the role of molecular markers in crop genomics is crucial for optimizing breeding strategies, conserving genetic resources, and developing resilient crop varieties to meet the demands of a growing population and ensure food security in a changing world.

Characterisation of the Cinnamomumparthenoxylon (Jack) Meisn (Lauraceae) transcriptome using Illumina paired-end sequencing and EST-SSR markers development for population genetics.

Cinnamomumparthenoxylon is an endemic and endangered species with significant economic and ecological value in Vietnam. A better understanding of the genetic architecture of the species will be useful when planning management and conservation. We aimed to characterize the transcriptome of C.parthenoxylon, develop novel molecular markers, and assess the genetic variability of the species. First, transcriptome sequencing of five trees (C.parthenoxylon) based on root, leaf, and stem tissues was performed for functional annotation analysis and development of novel molecular markers. The transcriptomes of C.parthenoxylon were analyzed via an Illumina HiSeqTM 4000 sequencing system. A total of 27,363,199 bases were generated for C.parthenoxylon. De novo assembly indicated that a total of 160,435 unigenes were generated (average length = 548.954 bp). The 51,691 unigenes were compared against different databases, i.e. COG, GO, KEGG, KOG, Pfam, Swiss-Prot, and NR for functional annotation. Furthermore, a total of 12,849 EST-SSRs were identified. Of the 134 primer pairs, 54 were randomly selected for testing, with 15 successfully amplified across nine populations of C.parthenoxylon. We uncovered medium levels of genetic diversity (PIC = 0.52, Na = 3.29, Ne = 2.18, P = 94.07%, Ho = 0.56 and He = 0.47) within the studied populations. The molecular variance was 10% among populations and low genetic differentiation (Fst = 0.06) indicated low gene flow (Nm = 2.16). A reduction in the population size of C.parthenoxylon was detected using BOTTLENECK (VP population). The structure analysis suggested two optimal genetic clusters related to gene flow among the populations. Analysis of molecular variance (AMOVA) revealed higher genetic variation within populations (90%) than among populations (10%). The UPGMA approach and DAPC divided the nine populations into three main clusters. Our findings revealed a significant fraction of the transcriptome sequences and these newlydeveloped novel EST-SSR markers are a very efficient tool for germplasm evaluation, genetic diversity and molecular marker-assisted selection in C.parthenoxylon. This study provides comprehensive genetic resources for the breeding and conservation of different varieties of C.parthenoxylon.

Customizable and stable multilocus chromosomal integration: a novel glucose-dependent selection system in Aureobasidium spp.

BackgroundNon-conventional yeasts hold significant potential as biorefinery cell factories for microbial bioproduction. Currently, gene editing systems used for these yeasts rely on antibiotic and auxotrophic selection mechanisms. However, the drawbacks of antibiotics, including high costs, environmental concerns, and the dissemination of resistance genes, make them unsuitable for large-scale industrial fermentation. For auxotrophic selection system, the engineered strains harboring auxotrophic marker genes are typically supplemented with complex nutrient-rich components instead of precisely defined synthetic media in large-scale industrial fermentations, thus lack selection pressure to ensure the stability of heterologous metabolic pathways. Therefore, it is a critical to explore alternative selection systems that can be adapted for large-scale industrial fermentation.ResultsHere, a novel glucose-dependent selection system was developed in a high pullulan-producing non-conventional strain A. melanogenum P16. The system comprised a glucose-deficient chassis cell Δpfk obtained through the knockout of the phosphofructokinase gene (PFK) and a series of chromosomal integration plasmids carrying a selection marker PFK controlled by different strength promoters. Utilizing the green fluorescent protein gene (GFP) as a reporter gene, this system achieved a 100% positive rate of transformation, and the chromosomal integration numbers of GFP showed an inverse relationship with promoter strength, with a customizable copy number ranging from 2 to 54. More importantly, the chromosomal integration numbers of target genes remained stable during successive inoculation and fermentation process, facilitated simply by using glucose as a cost-effective and environmental-friendly selectable molecule to maintain a constant and rigorous screening pressure. Moreover, this glucose-dependent selection system exhibited no significant effect on cell growth and product synthesis, and the glucose-deficient related selectable marker PFK has universal application potential in non-conventional yeasts.ConclusionHere, we have developed a novel glucose-dependent selection system to achieve customizable and stable multilocus chromosomal integration of target genes. Therefore, this study presents a promising new tool for genetic manipulation and strain enhancement in non-conventional yeasts, particularly tailored for industrial fermentation applications.

Diagnostic and Prognostic Markers for Pancreatitis and Pancreatic Ductal Adenocarcinoma.

Diagnostic markers are desperately needed for the early detection of pancreatic ductal adenocarcinoma (PDA). We describe sets of markers expressed in temporal order in mouse models during pancreatitis, PDA initiation and progression. Cell type specificity and the differential expression of PDA markers were identified by screening single cell (sc) RNAseq from tumor samples of a mouse model for PDA (KIC) at early and late stages of PDA progression compared to that of a normal pancreas. Candidate genes were identified from three sources: (1) an unsupervised screening of the genes preferentially expressed in mouse PDA tumors; (2) signaling pathways that drive PDA, including the Ras pathway, calcium signaling, and known cancer genes, or genes encoding proteins that were identified by differential mass spectrometry (MS) of mouse tumors and conditioned media from human cancer cell lines; and (3) genes whose expression is associated with poor or better prognoses (PAAD, oncolnc.org). The developmental progression of PDA was detected in the temporal order of gene expression in the cancer cells of the KIC mice. The earliest diagnostic markers were expressed in epithelial cancer cells in early-stage, but not late-stage, PDA tumors. Other early markers were expressed in the epithelium of both early- and late-state PDA tumors. Markers that were expressed somewhat later were first elevated in the epithelial cancer cells of the late-stage tumors, then in both epithelial and mesenchymal cells, or only in mesenchymal cells. Stromal markers were differentially expressed in early- and/or late-stage PDA neoplasia in fibroblast and hematopoietic cells (lymphocytes and/or macrophages) or broadly expressed in cancer and many stromal cell types. Pancreatitis is a risk factor for PDA in humans. Mouse models of pancreatitis, including caerulein treatment and the acinar-specific homozygous deletion of differentiation transcription factors (dTFs), were screened for the early expression of all PDA markers identified in the KIC neoplasia. Prognostic markers associated with a more rapid decline were identified and showed differential and cell-type-specific expression in PDA, predominately in late-stage epithelial and/or mesenchymal cancer cells. Select markers were validated by immunohistochemistry in mouse and human samples of a normal pancreas and those with early- and late-stage PDA. In total, we present 2165 individual diagnostic and prognostic markers for disease progression to be tested in humans from pancreatitis to late-stage PDA.

Whole-genome resequencing of grass carp (Ctenopharyngodon idella) for genome-wide association study on GCRV resistance

Grass carp (Ctenopharyngodon idella) is a highly productive freshwater fish species that annually contributes significant economic value to the aquaculture industry. However, its production is frequently affected by grass carp haemorrhagic disease (GCHD), which is caused by grass carp reovirus (GCRV). To elucidate the molecular mechanism of GCRV resistance in C. idella, whole-genome resequencing of both susceptible and resistant C. idella was conducted, yielding a total of 3,941,776 high quality single nucleotide polymorphisms (SNPs). A genome-wide association study (GWAS) then identified five significant SNPs, located on chromosomes 2, 3, 5, 6 and 10, associated with the trait of GCRV resistance. An integrative analysis of the GWAS and a previous transcriptome study was performed, and five common genes were identified, including b4galt2 (β-1,4-galactosyltransferase 2), kif2c (Kinesin family member 2C), jmjd8 (JmjC domain containing 8), selenom (Selenoprotein M), and znfx1 (Zinc finger NFX1-type containing 1). This study serves as a valuable basis for implementing marker-assisted selection (MAS) to improve GCRV resistance in C. idella breeding programs, and provides insight into developing methods to control the spread of GCHD.

Phenotypic and molecular characterization of pleiotropic resistance to rusts and powdery mildew in spring wheat

An attempt was undertaken to comprehend how phenotypic markers, leaf tip necrosis (LTN), and pseudo black chaff (PBC), are associated with pleiotropic resistance genes in wheat (Triticum spp.). Pleiotropic resistance to stripe rust (Puccinia striiformis f. sp. tritici), leaf rust (P. triticina), stem rust (P. graminis f. sp. tritici) and powdery mildew (Blumeria graminis f. sp. tritici) was evaluated in 57 Egyptian spring wheat cultivars at seedling stage in greenhouse and at adult plant stage in field during 2021/22 and 2022/23. At seedling stage, pleiotropic resistance rated moderately resistance (MR) was observed in five cultivars, Misr-3 (all three rusts), Misr-4 (stripe rust, leaf rust, powdery mildew), Giza-168 (leaf rust, stem rust, powdery mildew), Sakha-94 and Sids-13 (leaf rust, stem rust). At adult plant stage, Misr-4 outperformed all other evaluated cultivars, exhibiting a high level of pleiotropic adult plant resistance (PAPR) against all diseases investigated. High levels of PAPR were also noticed against the three rusts in nine cultivars, Misr-3, Sakha-93, Sakha-94, Sakha-95, Giza-156, Giza-168, Giza-171, Gemmeiza-10 and Sids-13, and moderate levels against powdery mildew in Misr-3, Sakha-95, Giza-156. Four PAPR genes were characterized in cultivars based on phenotypic and molecular markers. Phenotypic markers (LTN and PBC) were observed in 23 cultivars. Molecular marker csLV34 indicated the presence of Lr34/Yr18/Sr57/Pm38/Ltn1 in six cultivars, whilst marker Xwmc44 indicated that Lr46/Yr29/Sr58/Pm39/Ltn2 was present in five cultivars. Marker Xcfd71 indicated the presence of Lr67/Yr46/Sr55/Pm46/Ltn3 in 19 cultivars. Gene Sr2/Yr30/Lr27/Pbc was identified with marker csSr2 in three cultivars. A complete association between LTN and molecular markers was recorded, while a strong association (r = 0.73) was recorded between PBC and molecular markers. Findings demonstrated the reliability of phenotypic markers in predicting pleiotropic resistance in wheat, which would facilitate marker-assisted selection in breeding programs.

Lack of selection of antimalarial drug resistance markers after intermittent preventive treatment of schoolchildren (IPTsc) against malaria in northeastern Tanzania

ObjectiveIntermittent Preventive Treatment of schoolchildren (IPTsc) is recommended by WHO as a strategy to protect against malaria; to explore whether IPTsc with dihydroartemisinin-piperaquine (DP) or artesunate-amodiaquine (ASAQ) cause a selection of molecular markers in Plasmodium falciparum genes associated with resistance in children in seven schools in Tanga region, Tanzania. MethodsSNPs in P. falciparum genes Pfmdr1, Pfexo, Pfkelch13, and Pfcrt and copy number variations in Pfplasmepsin-2 and Pfmdr1 were assessed in samples collected at 12 months (visit 4, n=74) and 20 months (visit 6, n=364) after initiation of IPTsc and compared with the baseline prevalence (n=379). ResultsThe prevalence of Pfmdr1 N86 and Pfexo 415G was >99% and 0%, respectively without any temporal differences observed. The prevalence of Pfmdr1 184F changed significantly from baseline (52.2%) to visit 6 (64.6%) (χ2=6.11, P=0.013), but no differences were observed between the treatment arms (χ2=0.05, P=0.98). Finally, only minor differences in the amplification of Pfmdr1 were observed; from 10.2% at baseline to 16.7% at visit 6 (χ2=0.98, P=0.32). ConclusionsThe IPTsc strategy does not seem to pose a risk for the selection of markers associated with DP or ASAQ resistance. Continuously and timely surveillance of markers of antimalarial drug resistance is recommended.

A genome-wide association study reveals novel loci and candidate genes associated with plant height variation in Medicago sativa

BackgroundPlant height (PH) is an important agronomic trait influenced by a complex genetic network. However, the genetic basis for the variation in PH in Medicago sativa remains largely unknown. In this study, a comprehensive genome-wide association analysis was performed to identify genomic regions associated with PH using a diverse panel of 220 accessions of M. sativa worldwide.ResultsOur study identified eight novel single nucleotide polymorphisms (SNPs) significantly associated with PH evaluated in five environments, explaining 8.59–12.27% of the phenotypic variance. Among these SNPs, the favorable genotype of chr6__31716285 had a low frequency of 16.4%. Msa0882400, located proximal to this SNP, was annotated as phosphate transporter 3;1, and its role in regulating alfalfa PH was supported by transcriptome and candidate gene association analysis. In addition, 21 candidate genes were annotated within the associated regions that are involved in various biological processes related to plant growth and development.ConclusionsOur findings provide new molecular markers for marker-assisted selection in M. sativa breeding programs. Furthermore, this study enhances our understanding of the underlying genetic and molecular mechanisms governing PH variations in M. sativa.

Detection of minimal residual disease in acute myeloid leukemia: evaluating utility and challenges

This study discusses the importance of minimal residual disease (MRD) detection in acute myeloid leukemia (AML) patients using liquid biopsy and next-generation sequencing (NGS). AML prognosis is based on various factors, including genetic alterations. NGS has revealed the molecular complexity of AML and helped refine risk stratification and personalized therapies. The long-term survival rates for AML patients are low, and MRD assessment is crucial in predicting prognosis. Currently, the most common methods for MRD detection are flow cytometry and quantitative PCR, but NGS is being incorporated into clinical practice due to its ability to detect genomic aberrations in the majority of AML patients. Typically, bone marrow samples are used for MRD assessment, but using peripheral blood samples or liquid biopsies would be less invasive. Leukemia originates in the bone marrow, along with the cfDNA obtained from peripheral blood. This study aimed to assess the utility of cell-free DNA (cfDNA) from peripheral blood samples for MRD detection in AML patients. A cohort of 20 AML patients was analyzed using NGS, and a correlation between MRD assessment by cfDNA and circulating tumor cells (CTCs) in paired samples was observed. Furthermore, a higher tumor signal was detected in cfDNA compared to CTCs, indicating greater sensitivity. Challenges for the application of liquid biopsy in MRD assessment were discussed, including the selection of appropriate markers and the sensitivity of certain markers. This study emphasizes the potential of liquid biopsy using cfDNA for MRD detection in AML patients and highlights the need for further research in this area.

Fine mapping of QYrsv.swust-1BL for resistance to stripe rust in durum wheat Svevo.

Stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is a serious disease that affects wheat worldwide. There is a great need to develop cultivars with combinations of all-stage resistance (ASR) and adult-plant resistance (APR) genes for sustainable control of the disease. QYrsv.swust-1BL in the Italian durum wheat (Triticum turgidum ssp. durum) cultivar Svevo is effective against Pst races in China and Israel, and the gene has been previously mapped to the long arm of chromosome 1B. The gene is flanked by SNP (single nucleotide polymorphism) markers IWB5732 and IWB4839 (0.75 cM). In the present study, we used high-density 660K SNP array genotyping and the phenotypes of 137 recombinant inbred lines (RILs) to fine map the QYrsv.swust-1BL locus within a 1.066 Mb region in durum wheat Svevo (RefSeq Rel. 1.0) on chromosome arm 1BL. The identified 1.066 Mb region overlaps with a previously described map of Yr29/QYr.ucw-1BL, a stripe rust APR gene. Twenty-five candidate genes for QYrsv.swut-1BL were identified through comparing polymorphic genes within the 1.066 Mb region in the resistant cultivar. SNP markers were selected and converted to Kompetitive allele-specific polymerase chain reaction (KASP) markers. Five KASP markers based on SNP were validated in a F2 and F2:3 breeding population, providing further compelling evidence for the significant effects of QYrsv.swut-1BL. These markers should be useful in marker-assisted selection for incorporating Yr29/QYrsv.swust-1BL into new durum and common wheat cultivars for resistance to stripe rust.

Updates on Strawberry DNA Testing and Marker-Assisted Breeding at the University of Florida

ABSTRACT Developing new strawberry varieties with high level of disease resistance and superior fruit quality is critical for strawberry production. Despite the complexity of the octoploid strawberry genome, recent advances in genomics and DNA sequencing technologies have allowed us to identify causal sequence variants associated with specific phenotypes. These have enabled the development of new and improved DNA markers for integration in marker-assisted breeding (MAB) efforts. The use of MAB has significantly improved the efficiency of stacking loci responsible for disease resistance and fruit quality characteristics. The “Strawberry DNA Testing Handbook” is currently accessible in the Genome Database for Rosaceae and has been an essential resource for the national and international strawberry research community. Based on the information provided in the handbook, the UF strawberry breeding program implemented seedling selection from 2021 to 2023, retaining 25.3%, 19.2%, and 37.9% of seedlings for further evaluation in each respective year. In this report, we provide updates on DNA markers associated with disease resistance and fruit quality traits. Additionally, we demonstrate the practical implementation of these markers in high-throughput marker-assisted selection and their potential to enhance strawberry breeding.

Advanced strategies for single embryo selection in assisted human reproduction: A review of clinical practice and research methods

Among the primary objectives of contemporary assisted reproductive technology research are achieving the births of healthy singletons and improving overall fertility outcomes. Substantial advances have been made in refining the selection of single embryos for transfer, with the aim of maximizing the likelihood of successful implantation. The principal criterion for this selection is embryo morphology. Morphological evaluation systems are based on traditional parameters, including cell count and fragmentation, pronuclear morphology, cleavage rate, blastocyst formation, and various sequential embryonic assessments. To reduce the incidence of multiple pregnancies and to identify the single embryo with the highest potential for growth, invasive techniques such as preimplantation genetic screening are employed in in vitro fertilization clinics. However, new approaches have been suggested for clinical application that do not harm the embryo and that provide consistent, accurate results. Noninvasive technologies, such as time-lapse imaging and omics, leverage morphokinetic parameters and the byproducts of embryo metabolism, respectively, to identify noninvasive prognostic markers for competent single embryo selection. While these technologies have garnered considerable interest in the research community, they are not incorporated into routine clinical practice and still have substantial room for improvement. Currently, the most promising strategies involve integrating multiple methodologies, which together are anticipated to increase the likelihood of successful pregnancy.

Teaching old dogs new tricks: genetic engineering methanogens.

Methanogenic archaea, which are integral to global carbon and nitrogen cycling, currently face challenges in genetic manipulation due to unique physiology and limited genetic tools. This review provides a survey of current and past developments in the genetic engineering of methanogens, including selection and counterselection markers, reporter systems, shuttle vectors, mutagenesis methods, markerless genetic exchange, and gene expression control. This review discusses genetic tools and emphasizes challenges tied to tool scarcity for specific methanogenic species. Mutagenesis techniques for methanogens, including physicochemical, transposon-mediated, liposome-mediated mutagenesis, and natural transformation, are outlined, along with achievements and challenges. Markerless genetic exchange strategies, such as homologous recombination and CRISPR/Cas-mediated genome editing, are also detailed. Finally, the review concludes by examining the control of gene expression in methanogens. The information presented underscores the urgent need for refined genetic tools in archaeal research. Despite historical challenges, recent advancements, notably CRISPR-based systems, hold promise for overcoming obstacles, with implications for global health, agriculture, climate change, and environmental engineering. This comprehensive review aims to bridge existing gaps in the literature, guiding future research in the expanding field of archaeal genetic engineering.

CelFiE-ISH: a probabilistic model for multi-cell type deconvolution from single-molecule DNA methylation haplotypes

Deconvolution methods infer quantitative cell type estimates from bulk measurement of mixed samples including blood and tissue. DNA methylation sequencing measures multiple CpGs per read, but few existing deconvolution methods leverage this within-read information. We develop CelFiE-ISH, which extends an existing method (CelFiE) to use within-read haplotype information. CelFiE-ISH outperforms CelFiE and other existing methods, achieving 30% better accuracy and more sensitive detection of rare cell types. We also demonstrate the importance of marker selection and of tailoring markers for haplotype-aware methods. While here we use gold-standard short-read sequencing data, haplotype-aware methods will be well-suited for long-read sequencing.

Characterization of a Powdery Mildew Resistance Gene in Wheat Breeding Line Jingzi 102 Using Bulk Segregant RNA Sequencing.

Wheat (Triticum aestivum L.) is one of the most important crops worldwide. Powdery mildew caused by Blumeria graminis f. sp. tritici is a destructive disease threatening wheat yield and quality. The utilization of resistant genes and cultivars is considered the most economical, environmentally friendly, and effective method to control powdery mildew. Wheat breeding line Jingzi 102 was highly resistant to powdery mildew at both seedling and adult plant stages. Genetic analysis of F1, F2, and F2:3 populations of "Jingzi 102 × Shixin 828" showed that the resistance of Jingzi 102 against powdery mildew isolate E09 at the seedling stage was controlled by a single dominant gene, temporarily designated PmJZ. Using bulked segregant RNA sequencing combined with molecular markers analysis, PmJZ was located on the long arm of chromosome 2B and flanked by markers BJK695-1 and CIT02g-20 with the genetic distances of 1.2 and 0.5 centimorgan, respectively, corresponding to the bread wheat genome of Chinese Spring (International Wheat Genome Sequencing Consortium RefSeq v2.1) 703.8 to 707.6 Mb. PmJZ is most likely different from the documented Pm genes on chromosome 2BL based on their physical positions, molecular markers analysis, and resistance spectrum. Based on the gene annotation information, five genes related to disease resistance could be considered as the candidate genes of PmJZ. To accelerate the application of PmJZ, the flanking markers BJK695-1 and CIT02g-20 can serve for marker-assisted selection of PmJZ in wheat disease-resistance breeding.

CD52/FLAG and CD52/HA Fusion Proteins as Novel Magnetic Cell Selection Markers.

At present, the magnetic selection of genetically modified cells is mainly performed with surface markers naturally expressed by cells such as CD4, LNGFR (low affinity nerve growth factor receptor), and MHC class I molecule H-2Kk. The disadvantage of such markers is the possibility of their undesired and poorly predictable expression by unmodified cells before or after cell manipulation, which makes it essential to develop new surface markers that would not have such a drawback. Earlier, modified CD52 surface protein variants with embedded HA and FLAG epitope tags (CD52/FLAG and CD52/HA) were developed by the group of Dr. Mazurov for the fluorescent cell sorting of CRISPR-modified cells. In the current study, we tested whether these markers can be used for the magnetic selection of transduced cells. For this purpose, appropriate constructs were created in MigR1-based bicistronic retroviral vectors containing EGFP and DsRedExpress2 as fluorescent reporters. Cytometric analysis of the transduced NIH 3T3 cell populations after magnetic selection evaluated the efficiency of isolation and purity of the obtained populations, as well as the change in the median fluorescence intensity (MFI). The results of this study demonstrate that the surface markers CD52/FLAG and CD52/HA can be effectively used for magnetic cell selection, and their efficiencies are comparable to that of the commonly used LNGFR marker. At the same time, the significant advantage of these markers is the absence of HA and FLAG epitope sequences in cellular proteins, which rules out the spurious co-isolation of negative cells.

Ventral tegmental area interneurons revisited: GABA and glutamate projection neurons make local synapses.

The ventral tegmental area (VTA) contains projection neurons that release the neurotransmitters dopamine, GABA, and/or glutamate from distal synapses. VTA also contains GABA neurons that synapse locally on to VTA dopamine neurons, synapses widely credited to a population of so-called VTA interneurons. Interneurons in cortex, striatum, and elsewhere have well-defined morphological features, physiological properties, and molecular markers, but such features have not been clearly described in VTA. Indeed, there is scant evidence that local and distal synapses originate from separate populations of VTA GABA neurons. In this study we tested whether several markers expressed in non-dopamine VTA neurons are selective markers of interneurons, defined as neurons that synapse locally but not distally. Challenging previous assumptions, we found that VTA neurons genetically defined by expression of parvalbumin, somatostatin, neurotensin, or mu-opioid receptor project to known VTA targets including nucleus accumbens, ventral pallidum, lateral habenula, and prefrontal cortex. Moreover, we provide evidence that VTA GABA and glutamate projection neurons make functional inhibitory or excitatory synapses locally within VTA. These findings suggest that local collaterals of VTA projection neurons could mediate functions prior attributed to VTA interneurons. This study underscores the need for a refined understanding of VTA connectivity to explain how heterogeneous VTA circuits mediate diverse functions related to reward, motivation, or addiction.

Weighted single-step genome-wide association study for direct and maternal genetic effects associated with birth and weaning weights in sheep

Body weight is an important economic trait for sheep meat production, and its genetic improvement is considered one of the main goals in the sheep breeding program. Identifying genomic regions that are associated with growth-related traits accelerates the process of animal breeding through marker-assisted selection, which leads to increased response to selection. In this study, we conducted a weighted single-step genome-wide association study (WssGWAS) to identify potential candidate genes for direct and maternal genetic effects associated with birth weight (BW) and weaning weight (WW) in Baluchi sheep. The data used in this research included 13,408 birth and 13,170 weaning records collected at Abbas-Abad Baluchi Sheep Breeding Station, Mashhad-Iran. Genotypic data of 94 lambs genotyped by Illumina 50K SNP BeadChip for 54,241 markers were used. The proportion of variance explained by genomic windows was calculated by summing the variance of SNPs within 1 megabase (Mb). The top 10 window genomic regions explaining the highest percentages of additive and maternal genetic variances were selected as candidate window genomic regions associated with body weights. Our findings showed that for BW, the top-ranked genomic regions (1 Mb windows) explained 4.30 and 4.92% of the direct additive and maternal genetic variances, respectively. The direct additive genetic variance explained by the genomic window regions varied from 0.31 on chromosome 1 to 0.59 on chromosome 8. The highest (0.84%) and lowest (0.32%) maternal genetic variances were explained by genomic windows on chromosome 10 and 17, respectively. For WW, the top 10 genomic regions explained 6.38 and 5.76% of the direct additive and maternal genetic variances, respectively. The highest and lowest contribution of direct additive genetic variances were 1.37% and 0.42%, respectively, both explained by genomic regions on chromosome 2. For maternal effects on WW, the highest (1.38%) and lowest (0.41%) genetic variances were explained by genomic windows on chromosome 2. Further investigation of these regions identified several possible candidate genes associated with body weight. Gene ontology analysis using the DAVID database identified several functional terms, such as translation repressor activity, nucleic acid binding, dehydroascorbic acid transporter activity, growth factor activity and SH2 domain binding.

Identification of Resistance QTLs to Black Leaf Streak Disease (Due to Pseudocercospora fijiensis) in Diploid Bananas (Musa acuminata)

Black Leaf Streak Disease (BLSD), caused by the fungus Pseudocercospora fijiensis, is a recent pandemic and the most economically and environmentally important leaf disease of banana. To assist breeding of varieties with durable resistance to the rapidly evolving P. fijiensis, we used a diploid genitor ‘IDN 110’ with partial resistance to BLSD to search for QTLs. We assessed diploid progeny of 73 hybrids between ‘IDN 110’ and the diploid cultivar ‘Khai Nai On’, which is susceptible to BLSD. Hybrids were phenotyped with artificial inoculation under controlled conditions. This method allowed us to focus on resistance in the early stages of the interaction already identified as strongly influencing BLSD epidemiology. Progeny were genotyped by sequencing. As both parents are heterozygous for large reciprocal translocations, the distribution of recombination was assessed and revealed regions with low recombination rates. Fourteen non-overlapping QTLs of resistance to BLSD were identified of which four main QTLs from the ‘IDN110‘ parent, located on chromosomes 06, 07, 08, and 09, were shown to be of interest for marker-assisted selection. Genes that underline those four QTLs are discussed in the light of previous literature.

Identification and molecular mapping of resistance genes for adult-plant resistance to stripe rust in spring wheat germplasm line PI660076

Wheat is one of the major food crops worldwide. Stripe rust can cause a great loss of wheat yield, especially when the disease is prevalent. Chemical control not only causes the loss of resistance to stripe rust but also has a serious impact on the human body and environment. Therefore, the most economical measure to control wheat stripe rust is to cultivate resistant varieties. Rapid variation of stripe rust races often leads to rapid "loss" of resistance to stripe rust disease; therefore, breeders and researchers have to continuously explore new stripe rust resistance genes to provide new resistance sources for the rapid variation of stripe rust races. Previous studies have confirmed that PI660076, a spring wheat line, showed stripe rust resistance under natural conditions at the adult stage, which has great value in breeding programs. In this study, a recombinant inbred lines (RIL) population was constructed by crossing the wheat line PI660076 with the stripe rust-susceptible line AvS. Genotyping of the population was performed using a wheat 15 K SNP array. Three QTLs were identified using phenotypic data over four years across three environments. The resistance type of each QTL was determined by inoculating the RIL lines with single and homozygous QTL during the seedling and adult stages under controlled conditions. The all-stage resistance (ASR) QTL, QYr076.jaas-2A (flanked by SNP marker AX-11048464 and Kompetitive Allele-Specific PCR (KASP) marker KASP_4940) explained 7.13–16.58% and 6.95–7.25% of infection type (IT) and disease severity (DS), respectively. The adult-plant resistance (APR) resistance QTL, QYr076.jaas-4D.1 (flanked by KASP marker KASP_0795 and SNP marker AX-111567243,) explained 6.85–12.70% and 7.94–17.26% of IT and DS, respectively. The APR resistance QTL, QYr076.jaas-4D.2 flanked by KASP markers KASP_9130 and KASP_6535, explained 7.97–39.19% and 8.77–20.55% of the phenotypic variation in IT and DS, respectively. All the three QTLs are likely to be new. The obtained results lay a foundation for further utilization of the stripe rust-resistant line PI660076, as well as for fine mapping and molecular marker-assisted selection breeding.Graphical