Genetic mapping of the Andean anthracnose resistance gene present in the common bean cultivar BRSMG Realce.

The rajado seeded Andean bean (Phaseolus vulgaris L.) cultivar BRSMG Realce (striped seed coat) developed by Embrapa expressed a high level of anthracnose resistance, caused by Colletotrichum lindemuthianum, in field and greenhouse screenings. The main goal of this study was to evaluate the inheritance of anthracnose resistance in BRSMG Realce, map the resistance locus or major gene cluster previously named as Co-Realce, identify resistance-related positional genes, and analyze potential markers linked to the resistance allele. F2 plants derived from the cross BRSMG Realce × BRS FC104 (Mesoamerican) and from the cross BRSMG Realce × BRS Notável (Mesoamerican) were inoculated with the C. lindemuthianum races 475 and 81, respectively. The BRSMG Realce × BRS FC104 F2 population was also genotyped using the DArTseq technology. Crosses between BRSMG Realce and BAT 93 (Mesoamerican) were also conducted and resulting F2 plants were inoculated with the C. lindemuthianum races 65 and 1609, individually. The results shown that anthracnose resistance in BRSMG Realce is controlled by a single locus with complete dominance. A genetic map including 1,118 SNP markers was built and shown 78% of the markers mapped at a distances less than 5.0 cM, with a total genetic length of 4,473.4 cM. A major locus (Co-Realce) explaining 54.6% of the phenotypic variation of symptoms caused by the race 475 was identified in Pv04, flanked by the markers snp1327 and snp12782 and 4.48 cM apart each other. These SNPs are useful for marker-assisted selection, due to an estimated selection efficiency of 99.2%. The identified resistance allele segregates independently of the resistance allele Co-33 (Pv04) present in BAT 93. The mapped genomic region with 704,867 bp comprising 63 putative genes, 44 of which were related to the pathogen-host interaction. Based on all these results and evidence, anthracnose resistance in BRSMG Realce should be considered as monogenic, useful for breeding purpose. It is proposed that locus Co-Realce is unique and be provisionally designated as CoPv04R until be officially nominated in accordance with the rules established by the Bean Improvement Cooperative Genetics Committee.

Meckel Gruber and Joubert Syndrome Diagnosed Prenatally: Allelism between the Two Ciliopathies, Complexities of Mutation Types and Digenic Inheritance.

Background: Antenatally detected occipital encephalocele and polycystic kidneys are a common presentation of ciliopathies like Joubert syndrome and Meckel Gruber syndrome which have considerable genetic and phenotypic overlap. Case reports: We describe 3 cases of antenatally diagnosed occipital encephalocele and enlarged kidneys with fetal autopsy, histopathology & exome sequencing results. A novel nonsense variant in the CEP290 gene was reported in first case (Meckel syndrome). The second case shows the importance of fetal exome where the parents were carriers for 2 ciliopathy genes (TMEM138 & SDCCAG8). Diagnosis in this case was confirmed by fetal exome sequencing (Joubert syndrome). Multiexon deletion in TMEM67 and KIF14 present in trans was identified in the third case (Meckel syndrome), likely resulting in digenic inheritance. Conclusion: We report 2 cases of Meckel syndrome with a novel variant and multiexon deletion, and 1 case of Joubert syndrome which depicts the limitations of preconceptional carrier screening in ciliopathies due to overlapping phenotypes.

The morbid genome of ciliopathies: an update.

PURPOSE: Ciliopathies are highly heterogeneous clinical disorders of the primary cilium. We aim to characterize a large cohort of ciliopathies phenotypically and molecularly. METHODS: Detailed phenotypic and genomic analysis of patients with ciliopathies, and functional characterization of novel candidate genes. RESULTS: In this study, we describe 125 families with ciliopathies and show that deleterious variants in previously reported genes, including cryptic splicing variants, account for 87% of cases. Additionally, we further support a number of previously reported candidate genes (BBIP1, MAPKBP1, PDE6D, and WDPCP), and propose nine novel candidate genes (CCDC67, CCDC96, CCDC172, CEP295, FAM166B, LRRC34, TMEM17, TTC6, and TTC23), three of which (LRRC34, TTC6, and TTC23) are supported by functional assays that we performed on available patient-derived fibroblasts. From a phenotypic perspective, we expand the phenomenon of allelism that characterizes ciliopathies by describing novel associations including WDR19-related Stargardt disease and SCLT1- and CEP164-related Bardet-Biedl syndrome. CONCLUSION: In this cohort of phenotypically and molecularly characterized ciliopathies, we draw important lessons that inform the clinical management and the diagnostics of this class of disorders as well as their basic biology.

Pi5 and Pii Paired NLRs Are Functionally Exchangeable and Confer Similar Disease Resistance Specificity.

Effector-triggered immunity (ETI) is an effective layer of plant defense initiated upon recognition of avirulence (Avr) effectors from pathogens by cognate plant disease resistance (R) proteins. In rice, a large number of R genes have been characterized from various cultivars and have greatly contributed to breeding programs to improve resistance against the rice blast pathogen Magnaporthe oryzae. The extreme diversity of R gene repertoires is thought to be a result of co-evolutionary history between rice and its pathogens including M. oryzae. Here we show that Pii is an allele of Pi5 by DNA sequence characterization and complementation analysis. Pii-1 and Pii-2 cDNAs were cloned by reverse transcription polymerase chain reaction from the Pii -carrying cultivar Fujisaka5 . The complementation test in susceptible rice cultivar Dongjin demonstrated that the rice blast resistance mediated by Pii , similar to Pi5 , requires the presence of two nucleotide-binding leucine-rich repeat genes, Pii-1 and Pii-2 . Consistent with our hypothesis that Pi5 and Pii are functionally indistinguishable, the replacement of Pii-1 by Pi5-1 and Pii-2 by Pi5-2 , respectively, does not change the level of disease resistance to M. oryzae carrying AVR-Pii. Surprisingly, Exo70F3, required for Pii-mediated resistance, is dispensable for Pi5-mediated resistance. Based on our results, despite similarities observed between Pi5 and Pii, we hypothesize that Pi5 and Pii pairs require partially distinct mechanisms to function.

Epistasis confers resistance to Bt toxin Cry1Ac in the cotton bollworm.

Evolution of resistance by insect pests reduces the benefits of extensively cultivated transgenic crops that produce insecticidal proteins from Bacillus thuringiensis (Bt). Previous work showed that resistance to Bt toxin Cry1Ac, which is produced by transgenic cotton, can be conferred by mutations disrupting a cadherin protein that binds this Bt toxin in the larval midgut. However, the potential for epistatic interactions between the cadherin gene and other genes has received little attention. Here, we report evidence of epistasis conferring resistance to Cry1Ac in the cotton bollworm, Helicoverpa armigera, one of the world's most devastating crop pests. Resistance to Cry1Ac in strain LF256 originated from a field-captured male and was autosomal, recessive, and 220-fold relative to susceptible strain SCD. We conducted complementation tests for allelism by crossing LF256 with a strain in which resistance to Cry1Ac is conferred by a recessive allele at the cadherin locus HaCad. The resulting F1 offspring were resistant, suggesting that resistance to Cry1Ac in LF256 is also conferred by resistance alleles at this locus. However, the HaCad amino acid sequence in LF256 lacked insertions and deletions, and did not differ consistently between LF256 and a susceptible strain. In addition, most of the cadherin alleles in LF256 were not derived from the field-captured male. Moreover, Cry1Ac resistance was not genetically linked with the HaCad locus in LF256. Furthermore, LF256 and the susceptible strain were similar in levels of HaCad transcript, cadherin protein, and binding of Cry1Ac to cadherin. Overall, the results imply that epistasis between HaCad and an unknown second locus in LF256 yielded the observed resistance in the F1 progeny from the complementation test. The observed epistasis has important implications for interpreting results of the F1 screen used widely to monitor and analyze resistance, as well as the potential to accelerate evolution of resistance.

Identification and Validation of a New Source of Low Grain Cadmium Accumulation in Durum Wheat.

Cadmium (Cd) is a heavy metal that has no known biological function and is toxic for many living organisms. The maximum level of Cd concentration allowed in the international market for wheat grain is 0.2 mg kg-1 Because phenotyping for Cd uptake is expensive and time consuming, molecular markers associated with genes conferring low Cd uptake would expedite selection and lead to the development of durum cultivars with reduced Cd concentrations. Here, we identified single nucleotide polymorphisms (SNPs) associated with a novel low Cd uptake locus in the durum experimental line D041735, which has hexaploid common wheat in its pedigree. Genetic analysis revealed a single major QTL for Cd uptake on chromosome arm 5BL within a 0.3 cM interval flanked by SNP markers. Analysis of the intervening sequence revealed a gene with homology to an aluminum-induced protein as a candidate gene. Validation and allelism tests revealed that the low Cd uptake gene identified in this study is different from the closely linked Cdu1-B gene, which also resides on 5BL. This study therefore showed that the durum experimental line D041735 contains a novel low Cd uptake gene that was likely acquired from hexaploid wheat.

Genetic analysis of seedling resistance to crown rust in five diploid oat (Avena strigosa) accessions.

Crown rust, caused by Puccinia coronata Corda f. sp. avenae Eriks., is a serious menace in oats, for which resistance is an effective means of control. Wild diploid oat accessions are a source of novel resistances that first need to be characterised prior to introgression into locally adapted oat cultivars. A genetic analysis of resistance to crown rust was carried out in three diverse diploid oat accessions (CIav6956, CIav9020, PI292226) and two cultivars (Saia and Glabrota) of A. strigosa. A single major gene conditioning resistance to Australian crown rust pathotype (Pt) 0000-2 was identified in each of the three accessions. Allelism tests suggested that these genes are either the same, allelic, or tightly linked with less than 1 % recombination. Similarly, a single gene was identified in Glabrota, and possibly two genes in Saia; both cultivars previously reported to carry two and three crown rust resistance genes, respectively. The identified seedling resistance genes could be deployed in combination with other resistance gene(s) to enhance durability of resistance to crown rust in hexaploid oat. Current diploid and hexaploid linkage maps and molecular anchor markers (simple sequence repeat [SSR] and diversity array technology [DArT] markers) should facilitate their mapping and introgression into hexaploid oat.

Embryo-lethal phenotypes in early abp1 mutants are due to disruption of the neighboring BSM gene.

The Auxin Binding Protein1 (ABP1) has been identified based on its ability to bind auxin with high affinity and studied for a long time as a prime candidate for the extracellular auxin receptor responsible for mediating in particular the fast non-transcriptional auxin responses. However, the contradiction between the embryo-lethal phenotypes of the originally described Arabidopsis T-DNA insertional knock-out alleles ( abp1-1 and abp1-1s) and the wild type-like phenotypes of other recently described loss-of-function alleles ( abp1-c1 and abp1-TD1) questions the biological importance of ABP1 and relevance of the previous genetic studies. Here we show that there is no hidden copy of the ABP1 gene in the Arabidopsis genome but the embryo-lethal phenotypes of abp1-1 and abp1-1s alleles are very similar to the knock-out phenotypes of the neighboring gene, BELAYA SMERT ( BSM). Furthermore, the allelic complementation test between bsm and abp1 alleles shows that the embryo-lethality in the abp1-1 and abp1-1s alleles is caused by the off-target disruption of the BSM locus by the T-DNA insertions. This clarifies the controversy of different phenotypes among published abp1 knock-out alleles and asks for reflections on the developmental role of ABP1.

Characterization of novel Sorghum brown midrib mutants from an EMS-mutagenized population.

Reducing lignin concentration in lignocellulosic biomass can increase forage digestibility for ruminant livestock and saccharification yields of biomass for bioenergy. In sorghum (Sorghum bicolor (L.) Moench) and several other C4 grasses, brown midrib (bmr) mutants have been shown to reduce lignin concentration. Putative bmr mutants isolated from an EMS-mutagenized population were characterized and classified based on their leaf midrib phenotype and allelism tests with the previously described sorghum bmr mutants bmr2, bmr6, and bmr12. These tests resulted in the identification of additional alleles of bmr2, bmr6, and bmr12, and, in addition, six bmr mutants were identified that were not allelic to these previously described loci. Further allelism testing among these six bmr mutants showed that they represented four novel bmr loci. Based on this study, the number of bmr loci uncovered in sorghum has doubled. The impact of these lines on agronomic traits and lignocellulosic composition was assessed in a 2-yr field study. Overall, most of the identified bmr lines showed reduced lignin concentration of their biomass relative to wild-type (WT). Effects of the six new bmr mutants on enzymatic saccharification of lignocellulosic materials were determined, but the amount of glucose released from the stover was similar to WT in all cases. Like bmr2, bmr6, and bmr12, these mutants may affect monolignol biosynthesis and may be useful for bioenergy and forage improvement when stacked together or in combination with the three previously described bmr alleles.

Excisions of a defective transposable CACTA element (Tetu1) generate new alleles of a CYCLOIDEA-like gene of Helianthus annuus.

Tubular ray flower (turf) is a sunflower mutant that caught attention because it bears actinomorphic ray flowers, due to the presence of an active, although non-autonomous CACTA transposon (Tetu1) in the TCP domain of a CYCLOIDEA-like gene, HaCYC2c, a major regulator of sunflower floral symmetry. Here, we analyzed its excision rates in F3 population deriving from independent crosses of turf with common sunflower accessions. Our results suggest that the excision rate, ranging from 1.21 to 6.29%, depends on genetic background; moreover, the absence of somatic sectors in inflorescences of revertant individuals analyzed (182) and genetic analyses suggests a tight developmental control of Tetu1 excision, likely restricted to germinal cells. We individuate events of Tetu1 excision through molecular analysis that restore the wild type (WT) HaCYC2c allele, but even transposon excisions during which footprints are left. All mutations we detected occurred at the TCP basic motif and cause a change in ray flower phenotype. In particular, we selected five mutants with a one-to-four amino acid change that influence the capacity of reproductive organ development and ray flower corolla shaping (MUT-1, -2, -3, -4, -5). Revertant alleles not affecting turf phenotype (i.e. reading frame mutations) have also been identified (MUT-6). In all mutants, Real-time quantitative PCR (qPCR) experiments revealed variations of the steady state level of HaCYC2c mRNA. MUT-1 and MUT-4 showed a significant HaCYC2c down-regulation with respect to WT. A large variation within the biological replicates of MUT-2, MUT-3 and MUT-5 was detected and not significant differences in transcription levels between mutants and WT were observed. We detected low steady state level of HaCYC2c mRNA both in turf as in MUT-6. A three dimensional (3D) structure prediction tool let us predict an incorrect folding of the TCP protein already after a single amino acid deletion. This in turn is detectable as the restore of traits that are not peculiar of WT ray flowers, such as male fertility. Our analysis of an active TE sheds light on the TCP motif of the HaCYC2c gene and suggests that Tetu1 may be useful to obtain new natural mutants and for transposon tagging in different inbred lines of sunflower.

Modifiers of hearing impairment in humans and mice.

Lack of penetrance and variability of expression are common findings in nonsyndromic hearing loss with autosomal dominant mode of inheritance, but are also seen with recessive inheritance. Now we know that genotype cannot necessarily predict phenotype due to the complexity of the genome, the proteome interacting with the transcriptome, and the dynamically coupled systems that are involved. The contribution of genetic background to phenotypic diversity reflects the additive and interactive (epistasis) effects of multiple genes. Because, individual genes do not act alone but rather in concert with many other genes, it is not surprising that, modifier genes are common source of phenotypic variation in human populations. They can affect the phenotypic outcome of a given genotype by interacting in the same or in a parallel biological pathway as the disease gene. These modifier genes modulate penetrance, dominance, pleiotropy or expressivity in individuals with Mendelian traits and can also be exerted by influencing the severity, the penetrance, the age of onset and the progression of a disease. In this review, we focus on modifier genes that specifically affect hearing loss phenotypes in humans as well as those described in mice. We also include examples of digenic inheritance of deafness, because additive or interactive effects can also result from interaction between two mutant genes.

Genetic Analysis of Meloidogyne arenaria Race 1 Resistance in Tobacco.

Inheritance of resistance to the peanut root-knot nematode (Meloidogyne arenaria (Neal) Chitwood race 1) was investigated in the flue-cured tobacco cv. Speight G 28 and the breeding lines 81-RL-2K and SA 1214. The genetic relationship of this resistance in Speight G 28 to the resistance of the same cultivar to races 1 and 3 of M. incognita was also studied. Crosses were made between the root-knot nematode-susceptible flue-cured tobacco cv. NC 2326 and the three resistant genotypes. Parental, F1, F2 and backcross generations (BC1P1, BC1P2) were grown for each cross in randomized complete block designs with five replications in the greenhouse. Data indicated that resistance to M. arenaria race 1 in the three resistance sources is conditioned by a single dominant gene, but this resistance is partial compared to that for M. incognita races 1 and 3. Further, resistance to races 1 and 3 of M. incognita and resistance to M. arenaria race 1 in cv. Speight G 28 appear to be controlled by the same gene. These results, combined with the absence of segregation in the F2 populations of the crosses between resistant parents 81-RL-2K × SA 1214, 81-RL-2K × Speight G 28, and SA 1214 × Speight G 28, suggest allelism of resistance among these genotypes.