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1.
Artigo em Russo | MEDLINE | ID: mdl-34481439

RESUMO

Neuronal ceroid lipofuscinosis type 6 (NCL 6) is a rare progressive neurodegenerative disease that belongs to the group of lysosomal storage diseases. A clinical and genetic description of NCL 6 in a Yakut family was carried out. The proband and her sibling showed characteristic clinical signs, including myoclonic epilepsy, ataxia, psychomotor regression, dementia, and visual impairment. The onset of the disease in the age range from 3-4 years. The disease is caused by the frameshift mutation c.396dupT (p.Val133CysfsTer18) in exon 4 of the CLN6 in a homozygous state, which was detected using targeted next generation sequencing. Diagnosis of NCL is difficult due to the pronounced genetic heterogeneity of the disease, as well as the similarity with other hereditary metabolic diseases in clinical manifestations. The method of DNA diagnostics of NCL type 6 using NGS and direct sequencing according to Sanger has been introduced into the practice of medical genetic counseling.


Assuntos
Lipofuscinoses Ceroides Neuronais , Pré-Escolar , Feminino , Homozigoto , Humanos , Proteínas de Membrana/genética , Mutação , Lipofuscinoses Ceroides Neuronais/diagnóstico , Lipofuscinoses Ceroides Neuronais/genética
2.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 38(9): 873-876, 2021 Sep 10.
Artigo em Chinês | MEDLINE | ID: mdl-34487534

RESUMO

OBJECTIVE: To report the clinical manifestation and genetic characteristics of a child with Thiamine metabolism dysfunction syndrome 5. METHODS: Clinical data and genetic results were collected and analyzed. Peripheral blood samples of the child and their parents were collected for whole exome sequencing, and the functional effect of the variants on the TPK1 enzyme activity was verified by an in vitro assay. RESULTS: A four-year-old boy presented with preschool onset of ataxia were characterized. High-throughput sequencing identified a novel homozygous variant of TPK1 gene c.382G>A (p.Leu128Phe). His father and mother were both found carrying the variant. The variant protein showed a 30.9% reduction in TPK1 enzyme activity compared with the wildtype. CONCLUSION: A novel pathogenic variant has been identified in a boy with thiamine metabolic dysfunction syndrome type 5.


Assuntos
Testes Genéticos , Tiamina , Pré-Escolar , Homozigoto , Humanos , Masculino , Mutação , Sequenciamento Completo do Exoma
4.
Pan Afr Med J ; 39: 21, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34394812

RESUMO

Split-hand foot malformation (SHFM) is a clinically heterogeneous congenital limb defect affecting predominantly the central rays of hands and/or feet. The clinical expression varies in severity between patients as well between the limbs in the same individual. SHFM might be non-syndromic with limb-confined manifestations or syndromic with extra-limb manifestations. Isolated SHFM is a rare condition with an incidence of about 1 per 18,000 live born infants and accounts for 8-17 % of all limb malformations. To date, many chromosomal loci and genes have been described as associated with isolated SHFM, i.e., SHFM1 to 6. SHFM6 is one of the rarest forms of SHFM, and is caused by mutations in WNT10B gene. Less than ten pathogenic variants have been described. We have investigated a large consanguineous Moroccan family with three affected members showing feet malformations with or without split hand malformation phenotypes. Using an exome sequencing approach, we identified a homozygous nonsense variant p.Arg115* of WNT10B gene retaining thereby the diagnosis of SHFM6. This homozygous nonsense mutation identified by exome sequencing in a large family of split hand foot malformation highlights the importance of exome sequencing in genetically heterogeneous entities.


Assuntos
Deformidades Congênitas dos Membros/diagnóstico , Proteínas Proto-Oncogênicas/genética , Proteínas Wnt/genética , Criança , Códon sem Sentido , Exoma/genética , Feminino , Homozigoto , Humanos , Deformidades Congênitas dos Membros/genética , Deformidades Congênitas dos Membros/fisiopatologia , Marrocos
5.
Am J Hum Genet ; 108(9): 1692-1709, 2021 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-34375587

RESUMO

Kainate receptors (KARs) are glutamate-gated cation channels with diverse roles in the central nervous system. Bi-allelic loss of function of the KAR-encoding gene GRIK2 causes a nonsyndromic neurodevelopmental disorder (NDD) with intellectual disability and developmental delay as core features. The extent to which mono-allelic variants in GRIK2 also underlie NDDs is less understood because only a single individual has been reported previously. Here, we describe an additional eleven individuals with heterozygous de novo variants in GRIK2 causative for neurodevelopmental deficits that include intellectual disability. Five children harbored recurrent de novo variants (three encoding p.Thr660Lys and two p.Thr660Arg), and four children and one adult were homozygous for a previously reported variant (c.1969G>A [p.Ala657Thr]). Individuals with shared variants had some overlapping behavioral and neurological dysfunction, suggesting that the GRIK2 variants are likely pathogenic. Analogous mutations introduced into recombinant GluK2 KAR subunits at sites within the M3 transmembrane domain (encoding p.Ala657Thr, p.Thr660Lys, and p.Thr660Arg) and the M3-S2 linker domain (encoding p.Ile668Thr) had complex effects on functional properties and membrane localization of homomeric and heteromeric KARs. Both p.Thr660Lys and p.Thr660Arg mutant KARs exhibited markedly slowed gating kinetics, similar to p.Ala657Thr-containing receptors. Moreover, we observed emerging genotype-phenotype correlations, including the presence of severe epilepsy in individuals with the p.Thr660Lys variant and hypomyelination in individuals with either the p.Thr660Lys or p.Thr660Arg variant. Collectively, these results demonstrate that human GRIK2 variants predicted to alter channel function are causative for early childhood development disorders and further emphasize the importance of clarifying the role of KARs in early nervous system development.


Assuntos
Encéfalo/metabolismo , Deficiências do Desenvolvimento/genética , Epilepsia/genética , Deficiência Intelectual/genética , Mutação , Receptores de Ácido Caínico/genética , Adolescente , Adulto , Alelos , Encéfalo/diagnóstico por imagem , Encéfalo/patologia , Criança , Pré-Escolar , Deficiências do Desenvolvimento/diagnóstico por imagem , Deficiências do Desenvolvimento/metabolismo , Deficiências do Desenvolvimento/patologia , Epilepsia/diagnóstico por imagem , Epilepsia/metabolismo , Epilepsia/patologia , Potenciais Evocados/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Estudos de Associação Genética , Heterozigoto , Homozigoto , Humanos , Deficiência Intelectual/diagnóstico por imagem , Deficiência Intelectual/metabolismo , Deficiência Intelectual/patologia , Ativação do Canal Iônico , Masculino , Modelos Moleculares , Neurônios/metabolismo , Neurônios/patologia , Conformação Proteica , Receptores de Ácido Caínico/química , Receptores de Ácido Caínico/metabolismo
6.
Arch Oral Biol ; 130: 105246, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34454376

RESUMO

OBJECTIVE: To investigate whether null variants of Glutathione S-transferase Mu 1 (GSTM1) and GST Theta 1 (GSTT1) in infants and mothers, as well as maternal exposures to environmental factors, contribute to the risk of non-syndromic cleft lip with or without palate (NSCL/P) in a Mexican population. DESIGN: We performed a matched pair case-control study, including 98 cases and 98 controls and their mothers. Sociodemographic information and environmental exposures were collected by a questionnaire. Null variants of GSTM1 and GSTT1 were assessed by multiplex Polymerase Chain Reaction (PCR). Odds ratios (OR) and their 95 % confidence intervals (CI) were calculated to estimate risks. The interaction of genetic variables with smoking and adjusted ORs were evaluated by binary logistic regression. RESULTS: Homozygous null GSTM1 was associated with the risk of NSCL/P when present in mothers (OR = 2.45, 95 % CI 1.23-4.86) or infants (OR = 2.98, 95 % CI 1.45-6.14). A higher risk was also found when children carried the homozygous null GSTT1 (OR = 4.89, 95 % CI 2.42-9.87). In mothers, this variant showed a crude risk of 9.17 (95 % CI 3.95-21.29), which increased to OR = 13.81 (95 % CI 1.63-117.09) upon interaction with frequent passive smoking (5-7 days/week). Sociodemographic and other environmental exposures were not significantly associated with the risk of NSCL/P. CONCLUSIONS: Maternal and infant GSTT1 and GSTM1 homozygous null genotypes were associated with a higher risk of NSCL/P, and the results suggest an interaction of the maternal GSTT1-null/null genotype with frequent passive smoking.


Assuntos
Fissura Palatina , Glutationa Transferase , Estudos de Casos e Controles , Criança , Fissura Palatina/genética , Feminino , Predisposição Genética para Doença , Genótipo , Glutationa Transferase/genética , Homozigoto , Humanos , Exposição Materna , Polimorfismo Genético , Fatores de Risco , Deleção de Sequência
7.
Int J Mol Sci ; 22(15)2021 Aug 03.
Artigo em Inglês | MEDLINE | ID: mdl-34361096

RESUMO

Adenosine Deaminase 2 Deficiency (DADA2) syndrome is a rare monogenic disorder prevalently linked to recessive inherited loss of function mutations in the ADA2/CECR1 gene. It consists of an immune systemic disease including autoinflammatory vasculopathies, with a frequent onset at infancy/early childhood age. DADA2 syndrome encompasses pleiotropic manifestations such as stroke, systemic vasculitis, hematologic alterations, and immunodeficiency. Although skeletal abnormalities have been reported in patients with this disease, clear information about skeletal health, with appropriate biochemical-clinical characterization/management, its evolution over time and any appropriate clinical management is still insufficient. In this paper, after a general introduction shortly reviewing the pathophysiology of Ada2 enzymatic protein, its potential role in bone health, we describe a case study of two 27 year-old DADA2 monozygotic female twins exhibiting bone mineral density and bone turnover rate abnormalities over the years of their clinical follow-up.


Assuntos
Adenosina Desaminase/genética , Agamaglobulinemia/patologia , Predisposição Genética para Doença , Mutação em Linhagem Germinativa , Homozigoto , Peptídeos e Proteínas de Sinalização Intercelular/genética , Fenótipo , Imunodeficiência Combinada Severa/patologia , Adulto , Agamaglobulinemia/genética , Feminino , Humanos , Masculino , Linhagem , Imunodeficiência Combinada Severa/genética
8.
Nat Commun ; 12(1): 4228, 2021 07 09.
Artigo em Inglês | MEDLINE | ID: mdl-34244484

RESUMO

Homozygous deletion of methylthioadenosine phosphorylase (MTAP) in cancers such as glioblastoma represents a potentially targetable vulnerability. Homozygous MTAP-deleted cell lines in culture show elevation of MTAP's substrate metabolite, methylthioadenosine (MTA). High levels of MTA inhibit protein arginine methyltransferase 5 (PRMT5), which sensitizes MTAP-deleted cells to PRMT5 and methionine adenosyltransferase 2A (MAT2A) inhibition. While this concept has been extensively corroborated in vitro, the clinical relevance relies on exhibiting significant MTA accumulation in human glioblastoma. In this work, using comprehensive metabolomic profiling, we show that MTA secreted by MTAP-deleted cells in vitro results in high levels of extracellular MTA. We further demonstrate that homozygous MTAP-deleted primary glioblastoma tumors do not significantly accumulate MTA in vivo due to metabolism of MTA by MTAP-expressing stroma. These findings highlight metabolic discrepancies between in vitro models and primary human tumors that must be considered when developing strategies for precision therapies targeting glioblastoma with homozygous MTAP deletion.


Assuntos
Neoplasias Encefálicas/genética , Encéfalo/patologia , Desoxiadenosinas/metabolismo , Glioblastoma/genética , Purina-Núcleosídeo Fosforilase/deficiência , Tionucleosídeos/metabolismo , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Encéfalo/efeitos dos fármacos , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Meios de Cultivo Condicionados/metabolismo , Desoxiadenosinas/análise , Feminino , Secções Congeladas , Glioblastoma/tratamento farmacológico , Glioblastoma/patologia , Homozigoto , Humanos , Metabolômica , Metionina Adenosiltransferase/metabolismo , Terapia de Alvo Molecular/métodos , Medicina de Precisão/métodos , Proteína-Arginina N-Metiltransferases/metabolismo , Purina-Núcleosídeo Fosforilase/genética , Deleção de Sequência , Tionucleosídeos/análise , Ensaios Antitumorais Modelo de Xenoenxerto
9.
BMC Genomics ; 22(1): 538, 2021 Jul 13.
Artigo em Inglês | MEDLINE | ID: mdl-34256689

RESUMO

BACKGROUND: Although inbreeding caused by the mating of animals related through a recent common ancestor is expected to have more harmful effects on phenotypes than ancient inbreeding (old inbreeding), estimating these effects requires a clear definition of recent (new) and ancient (old) inbreeding. Several methods have been proposed to classify inbreeding using pedigree and genomic data. Unfortunately, these methods are largely based on heuristic criteria such as the number of generations from a common ancestor or length of runs of homozygosity (ROH) segments. To mitigate these deficiencies, this study aimed to develop a method to classify pedigree and genomic inbreeding into recent and ancient classes based on a grid search algorithm driven by the assumption that new inbreeding tends to have a more pronounced detrimental effect on traits. The proposed method was tested using a cattle population characterized by a deep pedigree. RESULTS: Effects of recent and ancient inbreeding were assessed on four growth traits (birth, weaning and yearling weights and average daily gain). Thresholds to classify inbreeding into recent and ancient classes were trait-specific and varied across traits and sources of information. Using pedigree information, inbreeding generated in the last 10 to 11 generations was considered as recent. When genomic information (ROH) was used, thresholds ranged between four to seven generations, indicating, in part, the ability of ROH segments to characterize the harmful effects of inbreeding in shorter periods of time. Nevertheless, using the proposed classification method, the discrimination between new and old inbreeding was less robust when ROH segments were used compared to pedigree. Using several model comparison criteria, the proposed approach was generally better than existing methods. Recent inbreeding appeared to be more harmful across the growth traits analyzed. However, both new and old inbreeding were found to be associated with decreased yearling weight and average daily gain. CONCLUSIONS: The proposed method provided a more objective quantitative approach for the classification of inbreeding. The proposed method detected a clear divergence in the effects of old and recent inbreeding using pedigree data and it was superior to existing methods for all analyzed traits. Using ROH data, the discrimination between old and recent inbreeding was less clear and the proposed method was superior to existing approaches for two out of the four analyzed traits. Deleterious effects of recent inbreeding were detected sooner (fewer generations) using genomic information than pedigree. Difference in the results using genomic and pedigree information could be due to the dissimilarity in the number of generations to a common ancestor. Additionally, the uncertainty associated with the identification of ROH segments and associated inbreeding could have an effect on the results. Potential biases in the estimation of inbreeding effects may occur when new and old inbreeding are discriminated based on arbitrary thresholds. To minimize the impact of inbreeding, mating designs should take the different inbreeding origins into consideration.


Assuntos
Endogamia , Polimorfismo de Nucleotídeo Único , Animais , Bovinos/genética , Genômica , Homozigoto , Linhagem , Fenótipo
10.
Methods Mol Biol ; 2288: 25-48, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270003

RESUMO

Doubled haploid (DH) technology produces strictly homozygous fertile plant thanks to doubling the chromosomes of a haploid embryo/seedling. Haploid embryos are derived from either male or female germ line cells and hold only half the number of chromosomes found in somatic plant tissues, albeit in a recombinant form due to meiotic genetic shuffling. DH production allows to rapidly fix these recombinant haploid genomes in the form of perfectly homozygous plants (inbred lines), which are produced in two rather than six or more generations. Thus, DH breeding enables fast evaluation of phenotypic traits on homogenous progeny. While for most crops haploid embryos are produced by costly and often genotype-dependent in vitro methods, for maize, two unique in planta systems are available to induce haploid embryos directly in the seed. Two "haploid inducer lines", identified from spontaneous maize mutants, are able to induce embryos of paternal or maternal origin. Although effortless crosses with lines of interest are sufficient to trigger haploid embryos, substantial improvements were necessary to bring DH technology to large scale production. They include the development of modern haploid inducer lines with high induction rates (8-12%), and methods to sort kernels with haploid embryos from the normal ones. Chromosome doubling represents also a crucial step in the DH process. Recent identification of genomic loci involved in spontaneous doubling opens up perspectives for a fully in planta DH pipeline in maize. Although discovered more than 60 years ago, maize haploid inducer lines still make headlines thanks to novel applications and findings. Indeed, maternal haploid induction was elegantly diverted to deliver genome editing machinery in germplasm recalcitrant to transformation techniques. The recent discovery of two molecular players controlling haploid induction allowed to revisit the mechanistic basis of maize maternal haploid induction and to successfully translate haploid induction ability to other crops.


Assuntos
Melhoramento Vegetal/métodos , Zea mays/genética , Cromossomos de Plantas/genética , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Cruzamentos Genéticos , Diploide , Edição de Genes , Genoma de Planta , Haploidia , Homozigoto , Vigor Híbrido , Modelos Genéticos , Biologia Molecular/métodos , Fenótipo , Sementes/genética , Sementes/crescimento & desenvolvimento , Zea mays/crescimento & desenvolvimento
11.
Methods Mol Biol ; 2288: 3-23, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270002

RESUMO

High frequency of albino plant formation in isolated microspore or anther cultures is a great problem limiting the possibility of their exploitation on a wider scale. It is highly inconvenient as androgenesis-based doubled haploid (DH) technology provides the simplest and shortest way to total homozygosity, highly valued by plant geneticists, biotechnologists and especially, plant breeders, and this phenomenon constitutes a serious limitation of these otherwise powerful tools. The genotype-dependent tendency toward albino plant formation is typical for many monocotyledonous plants, including cereals like wheat, barley, rice, triticale, oat and rye - the most important from the economical point of view. Despite many efforts, the precise mechanism underlying chlorophyll deficiency has not yet been elucidated. In this chapter, we review the data concerning molecular and physiological control over proper/disturbed chloroplast biogenesis, old hypotheses explaining the mechanism of chlorophyll deficiency, and recent studies which shed new light on this phenomenon.


Assuntos
Grão Comestível/crescimento & desenvolvimento , Grão Comestível/fisiologia , Pigmentação , Melhoramento Vegetal/métodos , Clorofila/deficiência , Clorofila/genética , Diploide , Grão Comestível/genética , Haploidia , Homozigoto , Modelos Biológicos , Biologia Molecular/métodos , Pigmentação/genética , Pigmentos Biológicos/deficiência , Pigmentos Biológicos/genética , Pólen/genética , Pólen/crescimento & desenvolvimento , Pólen/fisiologia , Regeneração/genética , Regeneração/fisiologia
12.
Methods Mol Biol ; 2288: 91-102, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270006

RESUMO

We describe the production of doubled haploids through anther culture in caraway. Induction conditions for the cultivation of donor plants, anther collection, composition of culture media, and physical induction conditions for embryogenesis have been described. As a result, responsive lines with numerous haploid embryo production were obtained, which after colchicine treatment became fertile. From a practical point of view, two doubled haploid populations are tested under field conditions.


Assuntos
Carum/crescimento & desenvolvimento , Carum/genética , Melhoramento Vegetal/métodos , Carum/fisiologia , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Meios de Cultura/química , Diploide , Esterases/análise , Flores/genética , Flores/crescimento & desenvolvimento , Haploidia , Homozigoto , Isoenzimas/análise , Biologia Molecular/métodos , Pólen/genética , Pólen/crescimento & desenvolvimento , Técnicas de Cultura de Tecidos
13.
Methods Mol Biol ; 2288: 49-72, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270004

RESUMO

Molecular markers are employed for doubled haploid (DH) technology by researchers and applied plant breeders in many crops. In the 1990s, isozymes and RFLPs were commonly used marker technologies to characterize DHs and were later replaced by PCR- based markers (e.g., RAPDs, AFLPs, ISSRs, SSRs) and today by SNPs. Markers are used for multiple purposes in DH production, that is, for the study of genes underlying haploid induction and confirming homozygous plants of gametophytic origin. Furthermore, they are tools for investigating segregation in DH populations and for mapping simple and complex traits using DHs. The deployment of DHs and markers for developing trait-linked markers are demonstrated with examples from rapeseed, wheat, and barley. Marker development for resistance to viruses derived from genetic resources and their use in, for example, pyramiding of resistance genes, are given as an example for the combination of DH-technology and marker development in research. Today, marker systems amenable to automation are frequently used in applied plant breeding. Practical examples are given from Lantmännen (LM) ( https://Lantmannen.com ) using large-scale genotyping for variety development based on SSRs and SNPs.


Assuntos
Produtos Agrícolas/genética , Melhoramento Vegetal/métodos , Brassica napus/genética , Produtos Agrícolas/virologia , DNA de Plantas/genética , Diploide , Resistência à Doença/genética , Genes de Plantas , Marcadores Genéticos , Haploidia , Homozigoto , Hordeum/genética , Isoenzimas/genética , Biologia Molecular/métodos , Doenças das Plantas/genética , Doenças das Plantas/virologia , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Polimorfismo de Nucleotídeo Único , Triticum/genética
14.
Methods Mol Biol ; 2288: 129-144, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270009

RESUMO

Rapeseed (Brassica napus) is one of the most important oilseed crops worldwide. It is also a model system to study the process of microspore embryogenesis, due to the high response of some B. napus lines, and to the refinements of the protocols. This chapter presents a protocol for the induction of haploid and DH embryos in B. napus through isolated microspore culture in two specific backgrounds widely used in DH research, the high response DH4079 line and the low response DH12075 line. We also present methods to identify the best phenological window to identify buds with microspores/pollen at the right developmental stage to induce this process. Methods to determine microspore/pollen viability and to check the ploidy by flow cytometry are also described.


Assuntos
Brassica napus/crescimento & desenvolvimento , Brassica napus/genética , Melhoramento Vegetal/métodos , Aclimatação/genética , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Meios de Cultura/química , Diploide , Citometria de Fluxo , Genótipo , Germinação/genética , Haploidia , Homozigoto , Biologia Molecular/métodos , Ploidias , Pólen/genética , Pólen/crescimento & desenvolvimento , Técnicas de Cultura de Tecidos
15.
Methods Mol Biol ; 2288: 113-126, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270008

RESUMO

Carrot is a vegetable of increasing economic importance. New hybrid cultivars are constantly required to meet the changing market needs. The application of anther culture significantly shortens the difficult and long-lasting breeding of carrot. We examined all the stages of the process of generating androgenic plants: induction of embryos in anther cultures, regeneration and acclimatization of produced plants, their evaluation, ploidy and homozygosity, and many other factors affecting their effectiveness. Every factor has been optimized by experimentally selecting the optimal level. As a result, a full protocol of producing homozygous plants using anther cultures was developed, which is presented in this chapter.


Assuntos
Daucus carota/crescimento & desenvolvimento , Daucus carota/genética , Melhoramento Vegetal/métodos , Aclimatação/genética , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Produtos Agrícolas/fisiologia , Meios de Cultura/química , Daucus carota/fisiologia , Flores/genética , Flores/crescimento & desenvolvimento , Heterozigoto , Homozigoto , Isoenzimas/análise , Biologia Molecular/métodos , Regeneração/genética , Técnicas de Cultura de Tecidos
16.
Methods Mol Biol ; 2288: 103-111, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270007

RESUMO

Doubled haploidy technology is a powerful tool to accelerate the breeding of new crop varieties. Protocols are not universal, as even species within the same family require a specific process. Here we describe methods for developing doubled haploids for fennel and dill, both Apiaceae species which are used for food, flavorings, and medicine.


Assuntos
Anethum graveolens/crescimento & desenvolvimento , Anethum graveolens/genética , Foeniculum/crescimento & desenvolvimento , Foeniculum/genética , Melhoramento Vegetal/métodos , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Meios de Cultura/química , Diploide , Haploidia , Homozigoto , Biologia Molecular/métodos , Pólen/genética , Pólen/crescimento & desenvolvimento , Técnicas de Cultura de Tecidos
17.
Methods Mol Biol ; 2288: 145-162, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270010

RESUMO

Culture of isolated microspores is a widely used method to obtain haploid and doubled haploid plants for many crop species. This protocol describes the steps necessary to obtain a large number of microspore derived embryos for pakchoi (Brassica rapa L. ssp. chinensis) and zicaitai (Brassica rapa L. ssp. сhinensis Hanelt var. purpuraria Kitam).


Assuntos
Brassica rapa/crescimento & desenvolvimento , Brassica rapa/genética , Melhoramento Vegetal/métodos , Brassica rapa/ultraestrutura , Cloroplastos/ultraestrutura , Cromossomos de Plantas/ultraestrutura , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Meios de Cultura/química , Diploide , Germinação/genética , Haploidia , Homozigoto , Microscopia de Fluorescência , Biologia Molecular/métodos , Ploidias , Pólen/genética , Pólen/crescimento & desenvolvimento , Técnicas de Cultura de Tecidos
18.
Methods Mol Biol ; 2288: 163-180, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270011

RESUMO

Brassica carinata, also known as Ethiopian or Abyssinian mustard, is a drought- and heat-tolerant oilseed with great potential as a dedicated industrial feedstock crop for use in biofuel and other bio-based applications. Doubled haploid technology, a system that allows for the rapid development of doubled haploid, completely homozygous plants through microspore embryogenesis, has been applied routinely in both B. carinata breeding and basic research. Here, we present a comprehensive isolated microspore culture protocol detailing the various steps involved in doubled haploid plant production for this species, from growing donor plants over harvesting flower buds and isolating, culturing and inducing microspores to regenerating doubled haploid embryos and plantlets.


Assuntos
Mostardeira/crescimento & desenvolvimento , Mostardeira/genética , Melhoramento Vegetal/métodos , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Meios de Cultura/química , Diploide , Haploidia , Homozigoto , Biologia Molecular/métodos , Ploidias , Pólen/genética , Pólen/crescimento & desenvolvimento , Pólen/ultraestrutura , Técnicas de Cultura de Tecidos
19.
Methods Mol Biol ; 2288: 181-199, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270012

RESUMO

The production of haploid and doubled haploid plants is a biotechnological tool that shortens the breeding process of new cultivars in many species. Doubled haploid plants are homozygous at every locus and they can be utilized as parents to produce F1 hybrids. In this chapter, we describe a protocol for the production of doubled haploid plants in Brassica rapa L. subsp. pekinensis using androgenesis induced by isolated microspore cultures.


Assuntos
Brassica rapa/crescimento & desenvolvimento , Brassica rapa/genética , Melhoramento Vegetal/métodos , Aclimatação/genética , Brassica rapa/fisiologia , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Produtos Agrícolas/fisiologia , Meios de Cultura/química , DNA de Plantas/genética , Diploide , Glucose-6-Fosfato Isomerase/genética , Haploidia , Homozigoto , Biologia Molecular/métodos , Pólen/genética , Pólen/crescimento & desenvolvimento , Reação em Cadeia da Polimerase , Regeneração/genética , Técnicas de Cultura de Tecidos
20.
Methods Mol Biol ; 2288: 201-216, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270013

RESUMO

Broccoli (Brassica olearecea var. italica) is a cole crop grown for its floral heads and stalks. It is rich in bioactive chemicals good for human health. Broccoli has been consumed as a vegetable since Roman times, but its production and consumption have increased significantly over the past few decades. Breeders try to develop new broccoli varieties with high yield, improved quality, and resistance to biotic and abiotic stresses. Almost all new broccoli varieties are F1 hybrids. Development of inbred broccoli lines that can be used as parents in hybrid production is a time-consuming and difficult process. Haploidization techniques can be utilized as a valuable support in broccoli breeding programs to speed up the production of genetically pure genotypes. Haploid plants of broccoli can be produced from immature male gametophytes via anther and microspore cultures with similar success rates. The most important parameters affecting the success of haploidization in broccoli are the genetic background (genotype) and the developmental stage of the microspores. Broccoli genotypes differ in their responses to androgenesis induction. The highest androgenesis response could be induced from microspores in late uninucleate and early binucleate stages. Recovery of diploid broccoli plants from haploids is possible via spontaneous and induced doubling. Doubled haploid (DH) broccoli lines are considered to be fully homozygous. Therefore, the production of DH lines is an alternative way to obtain pure inbred lines that can be utilized as parents in the development of new F1 hybrid varieties showing high levels of heterosis, high-quality heads, and uniform harvestable crop. We are using an anther culture-based haploid plant production system to develop DH broccoli lines in our broccoli breeding program. DH broccoli lines are produced from different genetic backgrounds within a year and handed to broccoli breeders.


Assuntos
Brassica/crescimento & desenvolvimento , Brassica/genética , Melhoramento Vegetal/métodos , Aclimatação/genética , Brassica/fisiologia , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Meios de Cultura/química , Diploide , Citometria de Fluxo , Flores/genética , Flores/crescimento & desenvolvimento , Haploidia , Homozigoto , Vigor Híbrido/genética , Biologia Molecular/métodos , Ploidias , Pólen/genética , Pólen/crescimento & desenvolvimento , Regeneração/genética , Técnicas de Cultura de Tecidos
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