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1.
Elife ; 102021 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-34596043

RESUMO

Evolutionary adaptation to a constant environment is driven by the accumulation of mutations which can have a range of unrealized pleiotropic effects in other environments. These pleiotropic consequences of adaptation can influence the emergence of specialists or generalists, and are critical for evolution in temporally or spatially fluctuating environments. While many experiments have examined the pleiotropic effects of adaptation at a snapshot in time, very few have observed the dynamics by which these effects emerge and evolve. Here, we propagated hundreds of diploid and haploid laboratory budding yeast populations in each of three environments, and then assayed their fitness in multiple environments over 1000 generations of evolution. We find that replicate populations evolved in the same condition share common patterns of pleiotropic effects across other environments, which emerge within the first several hundred generations of evolution. However, we also find dynamic and environment-specific variability within these trends: variability in pleiotropic effects tends to increase over time, with the extent of variability depending on the evolution environment. These results suggest shifting and overlapping contributions of chance and contingency to the pleiotropic effects of adaptation, which could influence evolutionary trajectories in complex environments that fluctuate across space and time.


Assuntos
Adaptação Biológica , Aptidão Genética , Pleiotropia Genética/fisiologia , Saccharomyces cerevisiae/fisiologia , Aclimatação , Diploide , Meio Ambiente , Haploidia , Saccharomyces cerevisiae/genética
2.
Elife ; 102021 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-34523420

RESUMO

Although studies of Saccharomyces cerevisiae have provided many insights into mutagenesis and DNA repair, most of this work has focused on a few laboratory strains. Much less is known about the phenotypic effects of natural variation within S. cerevisiae's DNA repair pathways. Here, we use natural polymorphisms to detect historical mutation spectrum differences among several wild and domesticated S. cerevisiae strains. To determine whether these differences are likely caused by genetic mutation rate modifiers, we use a modified fluctuation assay with a CAN1 reporter to measure de novo mutation rates and spectra in 16 of the analyzed strains. We measure a 10-fold range of mutation rates and identify two strains with distinctive mutation spectra. These strains, known as AEQ and AAR, come from the panel's 'Mosaic beer' clade and share an enrichment for C > A mutations that is also observed in rare variation segregating throughout the genomes of several Mosaic beer and Mixed origin strains. Both AEQ and AAR are haploid derivatives of the diploid natural isolate CBS 1782, whose rare polymorphisms are enriched for C > A as well, suggesting that the underlying mutator allele is likely active in nature. We use a plasmid complementation test to show that AAR and AEQ share a mutator allele in the DNA repair gene OGG1, which excises 8-oxoguanine lesions that can cause C > A mutations if left unrepaired.


Assuntos
Variação Genética , Mutação Puntual , Saccharomyces cerevisiae/genética , Alelos , Sistemas de Transporte de Aminoácidos Básicos/genética , Reparo do DNA , Diploide , Teste de Complementação Genética , Haploidia , Taxa de Mutação , Fenótipo , Saccharomyces cerevisiae/classificação , Proteínas de Saccharomyces cerevisiae/genética
3.
Elife ; 102021 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-34533136

RESUMO

Eukaryotic life cycles alternate between haploid and diploid phases and in phylogenetically diverse unicellular eukaryotes, expression of paralogous homeodomain genes in gametes primes the haploid-to-diploid transition. In the unicellular chlorophyte alga Chlamydomonas, KNOX and BELL TALE-homeodomain genes mediate this transition. We demonstrate that in the liverwort Marchantia polymorpha, paternal (sperm) expression of three of five phylogenetically diverse BELL genes, MpBELL234, and maternal (egg) expression of both MpKNOX1 and MpBELL34 mediate the haploid-to-diploid transition. Loss-of-function alleles of MpKNOX1 result in zygotic arrest, whereas a loss of either maternal or paternal MpBELL234 results in variable zygotic and early embryonic arrest. Expression of MpKNOX1 and MpBELL34 during diploid sporophyte development is consistent with a later role for these genes in patterning the sporophyte. These results indicate that the ancestral mechanism to activate diploid gene expression was retained in early diverging land plants and subsequently co-opted during evolution of the diploid sporophyte body.


Assuntos
Diploide , Células Germinativas Vegetais , Marchantia/genética , Genes de Plantas , Haploidia , Filogenia
4.
BMC Genomics ; 22(1): 699, 2021 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-34579651

RESUMO

BACKGROUND: Nutrition and cell size play an important role in the determination of caste differentiation in queen and worker of honeybees (Apis mellifera), whereas the haploid genome dominates the differentiation of drones. However, the effects of female developmental environment on the development of males remain unclear. In this study, young drone larvae were transferred into worker cells (WCs) or remained in drone cells (DCs) to rear drones. The drone larvae were also grafted into queen cells (QCs) for 48 h and then transplanted into drone cells until emerging. Morphological indexes and reproductive organs of these three types of newly emerged drones were measured. Newly emerged drones and third instar drone larvae from WCs, DCs and QCs were sequenced by RNA sequencing (RNA-Seq). RESULTS: The amount of food remaining in cells of the QC and WC groups was significantly different to that in the DC group at the early larval stage. Morphological results showed that newly emerged DC drones had bigger body sizes and more well-developed reproductive tissues than WC and QC drones, whereas the reproductive tissues of QC drones were larger than those of WC drones. Additionally, whole body gene expression results showed a clear difference among three groups. At larval stage there were 889, 1761 and 1927 significantly differentially expressed genes (DEGs) in WC/DC, QC/DC and WC/QC comparisons, respectively. The number of DEGs decreased in adult drones of these three comparisons [678 (WC/DC), 338 (QC/DC) and 518 (WC/QC)]. A high number of DEGs were involved in sex differentiation, growth, olfaction, vision, mammalian target of rapamycin (mTOR), Wnt signaling pathways, and other processes. CONCLUSIONS: This study demonstrated that the developmental environment of honeybee females can delay male development, which may serve as a model for understanding the regulation of sex differentiation and male development in social insects by environmental factors.


Assuntos
Diferenciação Sexual , Olfato , Animais , Abelhas/genética , Feminino , Haploidia , Larva/genética , Masculino , Análise de Sequência de RNA
5.
PLoS One ; 16(8): e0249505, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34343170

RESUMO

Assessment of genetic purity of parental inbred lines and their resultant F1 hybrids is an essential quality control check in maize hybrid breeding, variety release and seed production. In this study, genetic purity, parent-offspring relationship and diversity among the inbred lines were assessed using 92 single-nucleotide polymorphism (SNP) markers. A total of 188 maize genotypes, comprising of 26 inbred lines, four doubled haploid (DH) lines and 158 single-cross maize hybrids were investigated in this study using Kompetitive Allele Specific Polymerase Chain Reaction (KASP) genotyping assays. The bi-allelic data was analyzed for genetic purity and diversity parameters using GenAlex software. The SNP markers were highly polymorphic and 90% had polymorphic information content (PIC) values of > 0.3. Pairwise genetic distances among the lines ranged from 0.05 to 0.56, indicating a high level of dissimilarity among the inbred lines. A maximum genetic distance of (0.56) was observed between inbred lines CKDHL0089 and CML443 while the lowest (0.05) was between I-42 and I-40. The majority (67%) of the inbred lines studied were genetically pure with residual heterozygosity of <5%, while only 33% had heterozygosity levels of >5%. Inbred lines, which were not pure, require purification through further inbreeding. Cluster analysis partitioned the lines into three distinct genetic clusters with the potential to contribute new beneficial alleles to the maize breeding program. Out of the 68 hybrids (43%) that passed the parent-offspring test, seven hybrids namely; SCHP29, SCHP95, SCHP94, SCHP134, SCHP44, SCHP114 and SCHP126, were selected as potential candidates for further evaluation and release due to their outstanding yield performance.


Assuntos
Genes de Plantas/genética , Melhoramento Vegetal/métodos , Polimorfismo de Nucleotídeo Único/genética , Zea mays/genética , Alelos , Cromossomos de Plantas/genética , Variação Genética/genética , Haploidia , Hormônio do Crescimento Humano , Hibridização Genética/genética
6.
Sci Rep ; 11(1): 14472, 2021 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-34262060

RESUMO

In this study, clubroot resistance in the resynthesized European winter Brassica napus cv. 'Tosca' was introgressed into a Canadian spring canola line '11SR0099', which was then crossed with the clubroot susceptible spring line '12DH0001' to produce F1 seeds. The F1 plants were used to develop a doubled haploid (DH) mapping population. The parents and the DH lines were screened against 'old' pathotypes 2F, 3H, 5I, 6M and 8N of the clubroot pathogen, Plasmodiophora brassicae, as well as against the 'new' pathotypes 5X, 5L, 2B, 3A, 3D, 5G, 8E, 5C, 8J, 5K, 3O and 8P. Genotyping was conducted using a Brassica 15K SNP array. The clubroot screening showed that 'Tosca, '11SR0099' and the resistant DH lines were resistant to three (2F, 3H and 5I) of the five 'old' pathotypes and four (2B, 3O, 8E and 8P) of the 12 'new' pathotypes, while being moderately resistant to the 'old' pathotype 8N and the 'new' pathotypes 3D and 5G. 'Tosca' was susceptible to isolates representing pathotype 3A (the most common among the 'new' pathotypes) as well as pathotypes 6M, 5X, 5L, 5K and 8J. Linkage analysis and QTL mapping identified a ca. 0.88-0.95 Mb genomic region on the A03 chromosome of 'Tosca' as conferring resistance to pathotypes 2F, 3H, 5I, 2B, 3D, 5G, 8E, 3O and 8P. The identified QTL genomic region housed the CRk, Crr3 and CRd gene(s). However, the susceptibility of 'Tosca' to most of the common virulent pathotypes makes it unattractive as a sole CR donor in the breeding of commercial canola varieties in western Canada.


Assuntos
Brassica napus/genética , Brassica napus/microbiologia , Doenças das Plantas/microbiologia , Plasmodioforídeos/patogenicidade , Alberta , Resistência à Doença/genética , Ligação Genética , Haploidia , Melhoramento Vegetal , Doenças das Plantas/genética , Plasmodioforídeos/isolamento & purificação , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas
7.
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
8.
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
9.
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
10.
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
11.
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
12.
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
13.
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
14.
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
15.
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
16.
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
17.
Methods Mol Biol ; 2288: 217-232, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270014

RESUMO

Here, we describe the first protocol of European radish (Raphanus sativus L. subsp. sativus convar. radicula) for obtaining doubled haploid plants through in vitro microspore culture, in which the full cycle of doubled haploid formation was successfully achieved. Using this protocol, a yield of up to eight embryoids per Petri dish can be obtained. Effectiveness of this protocol was confirmed for several genotypes of European radish.


Assuntos
Melhoramento Vegetal/métodos , Raphanus/crescimento & desenvolvimento , Raphanus/genética , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Produtos Agrícolas/fisiologia , Meios de Cultura/química , Diploide , Corantes Fluorescentes , Genótipo , Haploidia , Homozigoto , Indóis , Biologia Molecular/métodos , Ploidias , Pólen/genética , Pólen/crescimento & desenvolvimento , Raphanus/fisiologia , Regeneração/genética , Coloração e Rotulagem , Técnicas de Cultura de Tecidos
18.
Methods Mol Biol ; 2288: 235-250, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270015

RESUMO

Eggplant is one of the five important, worldwide-distributed solanaceous crops. The use of anther culture technology to produce pure, 100% homozygous doubled haploid lines for hybrid seed production is possible since 1982, where the first protocol of wide application to different eggplant materials was published. From then on, different improvements and adaptations to different materials have been made. In parallel, protocols to implement isolated microspore culture technology in eggplant have been developed principally in the last decade, which opens the door for a more efficient DH production in this species. In this chapter, two protocols, one for anther and other for isolated microspore culture in eggplant, are described. Some steps and materials are common to both approaches. A detailed description of each step from is provided.


Assuntos
Melhoramento Vegetal/métodos , Solanum melongena/crescimento & desenvolvimento , Solanum melongena/genética , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Produtos Agrícolas/fisiologia , Meios de Cultura/química , Diploide , Flores/genética , Flores/crescimento & desenvolvimento , Corantes Fluorescentes , Haploidia , Homozigoto , Indóis , Biologia Molecular/métodos , Ploidias , Pólen/genética , Pólen/crescimento & desenvolvimento , Regeneração/genética , Solanum melongena/fisiologia , Coloração e Rotulagem , Técnicas de Cultura de Tecidos
19.
Methods Mol Biol ; 2288: 251-266, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270016

RESUMO

The shed-microspore culture technique is an alternative sub-method combining anther and isolated microspore culture to induce microspore embryogenesis. Recently, its effective use in different types of peppers has drawn attention, because it has a higher embryo yield potential compared to anther culture and is more practical than isolated microspore culture. In this chapter, a stepwise protocol for shed-microspore culture of ornamental pepper is described. This protocol includes the steps of donor plant growth conditions, the choice of suitable flower buds based on DAPI staining of microspores, application of a cold pretreatment to flower buds, surface sterilization of the buds, shed-microspore culture of anthers, stress treatments, regeneration of androgenic in vitro plantlets, their acclimatization and ploidy analysis, and in vivo chromosome doubling of the haploid plants.


Assuntos
Capsicum/crescimento & desenvolvimento , Capsicum/genética , Melhoramento Vegetal/métodos , Pólen/crescimento & desenvolvimento , Pólen/genética , Capsicum/fisiologia , Produtos Agrícolas/genética , Produtos Agrícolas/crescimento & desenvolvimento , Produtos Agrícolas/fisiologia , Meios de Cultura/química , Diploide , Flores/genética , Flores/crescimento & desenvolvimento , Corantes Fluorescentes , Haploidia , Homozigoto , Indóis , Biologia Molecular/métodos , Ploidias , Regeneração/genética , Coloração e Rotulagem , Técnicas de Cultura de Tecidos
20.
Methods Mol Biol ; 2288: 267-278, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34270017

RESUMO

An efficient anther culture on double-layered media to produce doubled haploid (DH) plants in pepper (Capsicum annuum) was clearly shown to outperformed other techniques such as anther culture on solid medium and direct isolated microspore culture on liquid medium. It was even used for DH production in a cayenne type of hot pepper which was previously known as less responsive or even more recalcitrant to androgenesis than sweet bell pepper. Indeed, anther culture on double-layered media has been routinely used to produce DH plants on broad genotypes of C. annuum as parental candidates to develop hybrid varieties. The step-by-step protocol of pepper anther culture on double-layered media, we hereby present in detail, includes the growth of donor plants, the use of flower buds as anther source, flower bud disinfection, anther isolation, anther culture and incubation processes, embryo germination and plant acclimatization process, and transplanting of plants to the soil-compost medium in pots.


Assuntos
Capsicum/crescimento & desenvolvimento , Flores/crescimento & desenvolvimento , Germinação , Técnicas de Embriogênese Somática de Plantas/métodos , Técnicas de Cultura de Tecidos/métodos , Capsicum/genética , Meios de Cultura , Flores/genética , Haploidia
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