玉米花药培养和单倍体育种的研究新进展

利用花药培养获得单倍体,从而加速育种进程,是一顶新兴的生物技术,目前在玉米育种中广泛应用,本文综合近几年来国内外玉米的花药培养、单位体育种以及基因工程等方面的研究进展,重点对影响玉米花药培养效率的诸多因素进行了详细论述,并讨论了利用单7保体植株进行基因转导的潜力。     

玉米花药培养的研究初报

玉米是我国主要的粮食作物之一。利用自交系杂交以获得强大的杂种优势,是玉米育种工作中获得增产最有效的方法。但要获得自交系,按常规育种必须连续进行4—6年的自交,而且不能保证其纯合性。如能利用花药培养这一新技术来诱导玉米单倍体,便可作为纯合二倍体的材料,并可大大缩短培育自交系年限,迅速淘汰有害的隐性基因。1975年我国科学工作者首次获得玉米花粉植株,在玉米单倍体育种的征途上迈出了可喜的一步。我们从1973年起开展这一工作。1975年,我们两个单位协作,使工作有了初步进展。几     

番茄花药培养研究进展及展望

在番茄育种中,利用花药培养产生单倍体植株,可以比常规育种方案较短时期固定和分析杂种优势的新遗传组合,建立纯合番茄品系,本文综述了国内外番茄花药培养研究概况及进展,并提供了展望。     

陆地棉花药愈伤组织的诱导和继代培养

花药培养是自６０年代以来发展起来的一项生物技术,旨在诱导出单倍体花粉植株,应用于育种实践和基础研究〔１〕。以单倍体植株为基础的单倍体育种被认为在以下几个方面具有优越性：第一,缩短育种年限;第二,排除杂种优势对后代选择的干扰〔２,３〕。此外,花药培养在...     

烟草花粉单倍体育种的几个遗传学问题研究

烟草花药培养成功之后,国内外育种工作者用这一方法在烟草育种上做出了贡献,并在培养技术、染色体加倍方法、单倍体和加倍单倍体特性、花粉单倍体植株性状遗传变异及生活力等方面都进行了不同程度的研究。然而对花粉单倍体植株的遗传学问题,目前报道的为数不多,尚有不少有待进一步探讨的问题。     

单倍体小麦染色体加倍的研究

从七十年代利用花药培养技术获得单倍体植株以来,单倍体植物在育种应用上的潜力日益显示出来。但是这种潜力只有使单倍体植物加倍成为纯合二倍体才有可能发挥。因此,对于单倍体植物的染色体加倍,许多植物育种工作者进行过多方面的试验研究,提出过多种加倍方法,其中秋水仙碱法应用最为广泛,但加倍成功率并不理想,植株死亡率也较高。近年来有人用秋水仙碱和DMSO结合处理单倍体植株,加倍效果显著提高。就小麦来说,最高成功率有达9.0％     

甘蔗花粉植株诱导成功

甘蔗现有品种基本上都是远缘的种间杂种,杂合 程度很高。若能通过花药培养获得双单倍体植株,就 为获得甘蔗的纯系建立了极其有效的方法。甘蔗的纯 系不仅可用于遗传学研究,还可直接用于选配优良的 杂交组合,在甘蔗育种业中充分利用杂种优势,可大 大提高蔗糖的产量。甘蔗花药培养方法的建立为甘蔗 单倍体诱变开辟了广阔的前景,可使诱变育种效率大 大提高。美国在夏威夷进行了多年花药培养的研究^[2], 菲律宾也进行了这方面的研究甘蔗现有品种基本上都是远缘的种间杂种,杂合 程度很高。若能通过花药培养获得双单倍体植株,就 为获得甘蔗的纯系建立了极其有效的方法。甘蔗的纯 系不仅可用于遗传学研究,还可直接用于选配优良的 杂交组合,在甘蔗育种业中充分利用杂种优势,可大 大提高蔗糖的产量。甘蔗花药培养方法的建立为甘蔗 单倍体诱变开辟了广阔的前景,可使诱变育种效率大 大提高。美国在夏威夷进行了多年花药培养的研究^[2], 菲律宾也进行了这方面的研究^[2],但仅得到来源于花 粉的多细胞球。我国台湾省曾进行了甘蔗花药培养的 研究,但尚未见获得花粉小植株的报道。,但仅得到来源于花 粉的多细胞球。我国台湾省曾进行了甘蔗花药培养的 研究,但尚未见获得花粉小植株的报道。     

植物单倍体诱导技术发展与创新

单倍体育种是培育作物新品种的主要育种技术之一,提高单倍体诱导频率和简化诱导程序是单倍体育种技术的关键。随着单倍体诱导技术的发展与改进,单倍体育种技术已被广泛应用于许多重要植物的育种研究中,展现出基因纯合快速、育种年限缩短、育种效率提高等优势。单倍体诱导技术与杂交育种、诱变育种、反向育种和分子标记辅助选择育种等技术相结合,在作物品种改良上的作用更加显著。单倍体和双单倍体在遗传群体构建、基因功能鉴定、转基因研究、细胞学研究等方面具有重要应用价值。本文从单倍体诱导技术、单倍体和双单倍体应用等方面综述了植物单倍体诱导技术的发展,尤其是近年来利用基因组编辑技术创制主要作物单倍体诱导系的进展,并分析了目前研究中存在的问题和今后的发展方向,以期促进单倍体诱导技术尤其是利用基因编辑创造诱导系技术在作物育种中的应用。     

植物遗传工程 八、在实际应用方面的潜力

在前一讲里,我们谈到了植物遗传工程在理论问题方面的研究进展。在这里我着重讲讲这一个新的领域在较近期内对育种实践上可能应用的潜力。 我们在前几次曾经谈到花药培养,通过单倍体育种,加快育种过程;也谈到了把外源基因注入植物体内的实验。今天着重谈谈育种中的另一个重要问题——配合力(cambining ability)。     

玉米花粉植株生理特性的初步研究

通过花药培养,产生单倍体植株,有可能成为获得玉米纯系的一条捷径。因此玉米花药培养的研究引起了国内外遗传育种工作者的普遍关注。我国自1975年首次培养成功以来,已有20多个单位成功地获得了玉米花粉植株,得苗3000株以上,但存在着试管苗移栽成活率不到20％,自交结实率不到2％的问题。为使这项研究成果尽快应用于育种实践,对移栽     

Rapid attainment of a doubled haploid line from transgenic maize (<Emphasis Type="Italic">Zea mays</Emphasis> L.) plants by means of anther culture

Summary We present a strategy for establishing a transgenic doubled haploid maize line from heterozygous transgenic material by means of anther culture. Compared to conventional inbreeding, the in vitro androgenesis technique enables a faster generation of virtually fully homozygous lines. Since the androgenic response is highly genotype-dependent, we crossed transgenic, non-androgenic plants carrying a herbicide resistance marker gene (pat, encoding for phosphinothricin acetyl transferase) with a highly androgenic genotype. The transgenic progenies were used as donor plants for anther culture. One transgenic and three non-transgenic doubled haploid lines have been established within approximately 1 yr. The homozygosity of all four doubled haploid lines was tested by analysis of simple sequence repeat (SSR) markers at 19 different loci. Polymorphisms were found between the lines but not within the lines indicating the homozygous nature of the entire plant genome gained by anther culture. Southern blot analysis revealed that the transgenic donor plants and their doubled haploid progeny exhibited the same integration pattern of the pat gene. No segregation of the herbicide resistance trait has been observed among the progeny of the transgenic doubled haploid line.     

Doubled haploid production in Flax (Linum usitatissimum L.)

There is a requirement of haploid and double haploid material and homozygous lines for cell culture studies and breeding in flax. Anther culture is currently the most successful method producing doubled haploid lines in flax. Recently, ovary culture was also described as a good source of doubled haploids. In this review we focus on tissue and plants regeneration using anther culture, and cultivation of ovaries containing unfertilized ovules. The effect of genotype, physiological status of donor plants, donor material pre-treatment and cultivation conditions for flax anthers and ovaries is discussed here. The process of plant regeneration from anther and ovary derived calli is also in the focus of this review. Attention is paid to the ploidy level of regenerated tissue and to the use of molecular markers for determining of gametic origin of flax plants derived from anther and ovary cultures. Finally, some future prospects on the use of doubled haploids in flax biotechnology are outlined here.     

Efficient production of doubled haploid plants through colchicine treatment of anther-derived maize callus

Summary A chromosome doubling technique, involving colchicine treatment of an embryogenic, haploid callus line of maize (Zea mays L., derived through anther culture), was evaluated. Two colchicine levels (0.025% and 0.05%) and three treatment durations (24, 48, and 72 h) were used and compared to untreated controls. Chromosome counts and seed recovery from regenerated plants were determined. No doubled haploid plants were regenerated from calli without colchicine treatment. After treatment with colchicine for 24 h, the callus tissue regenerated about 50% doubled haploid plants. All of the plants regenerated from the calli treated with colchicine for 72 h were doubled haploids, except for a few tetraploid plants. No significant difference in chromosome doubling was observed between the two colchicine levels. Most of the doubled haploid plants produced viable pollen and a total of 107 of 136 doubled haploid plants produced from 1 to 256 seeds. Less extensive studies with two other genotypes gave similar results. These results demonstrate that colchicine treatment of haploid callus tissue can be a very effective and relatively easy method of obtaining a high frequency of doubled haploid plants through anther culture.     

甜(辣)椒单倍体培养研究进展

介绍了甜(辣)椒花药培养和游离小孢子培养的研究概况,花药培养应用相对成熟,游离小孢子培养尚未获得突破性进展。对影响花药培养的各关键因素(包括材料基因型、供体植株生长状态、小孢子发育时期、培养基、培养方法、变温处理、培养条件等)进行了综述,并讨论了甜(辣)椒单倍体培养存在的问题和进一步研究方向。     

The agronomic performance of anther culture derived plants of barley produced via pollen embryogenesis

Anther culture was used to generate microspore-derived doubled haploid (DH) plants from four spring barley crosses. The culture medium used contained maltose as the sole carbohydrate source and the mode of plantlet regeneration was mainly via pollen embryogenesis. Both haploid and spontaneously doubled regenerants were produced and the doubled haploids were compared to recom-binant inbred lines generated by several rounds of selfing (single seed descent). Parental, DH and single seed descent (SSD) lines were grown in randomised, replicated field trials and the samples were scored for a range of agronomic traits. The mean performance and phenotypic distribution of the DH and SSD samples were similar and there was little evidence to support the conclusion that anther culture derived lines exhibit a reduction in vigour. Where significant differences were detected between groups these were mainly confined to crosses which were segregating for the denso dwarfing gene. The differential transmission of particular regions of the barley genome may therefore influence and confound the expression of agronomic traits in DH populations. This is the first report of the agronomic performance of anther culture lines produced via pollen embryogenesis and the results are discussed in relation to the exploitation of anther culture technology in barley breeding.     

Production of doubled haploids in durum wheat (Triticum turgidum L.) through isolated microspore culture

The objective of this work was to produce doubled haploid plants from durum wheat through the induction of androgenesis. A microspore culture technique was developed and used to produce fertile doubled haploid plants of agronomic interest. Five cultivars, one selected line, plus a collection of 20 F₁ crosses between different genotypes of high breeding value were used. Studies on several factors such as pre-treatments and media components were carried out in order to develop a protocol to regenerate green haploid plantlets. Anthers were pre-treated in 0.7 M mannitol. Microspores, from anther maceration, were plated on a C₁₇ induction culture medium with ovary co-culture. The optimum regeneration medium J25–8 was used. From 35 microspore isolations, 407 green plantlets were obtained. With this technique mature embryos were obtained. Green plants were regenerated from all genotypes used and approximately 67% of them were spontaneously doubled haploids. Some haploids and a very few polyploids plants were obtained. From the 407 plants, 275 were completely fertile and gave enough seeds to be assayed in the field. This protocol could be used complementary to or instead of the intergeneric crossing with maize as an economically feasible method to obtain doubled haploids from most durum wheat genotypes.     

Stimulation of embryogenesis and haploid production in Brassica campestris anther cultures by elevated temperature treatments

Summary Culture of Brassica campestris anthers at 35°C for one or three days prior to culture at 25°C significantly stimulated the yield of microspore-derived embryos. More than 100 plants were regenerated from cultured embryos and haploids were identified amongst them. The haploid frequency was greater than 70% if all small-flowered sterile plants were considered to be haploid. The yield of microspore-derived plants in B. campestris is approaching the level where anther culture may be utilized as a practical breeding tool.     

A simple wheat haploid and doubled haploid production system using anther culture

Summary Wheat (Triticum aestivum L.) haploids and doubled haploids have been used in breeding programs and genetic studies. Wheat haploids and doubled haploids via anther culture are usually produced by a multiple step culture procedure. We improved a wheat haploid and doubled haploid production system via anther culture in which plants are produced from microspore-derived embryos using one medium and one culture environment. In the improved protocol, tillers of donor plants were pretreated at 4°C for 1–2 wk before anthers were plated on a modified 85D12 basal medium with phenylacetic acid (PAA) and zeatin and cultured at 30°C with a 12-h daylength (43 μEs⁻¹m⁻²) in an incubator. Microspore-derived embryos developed in 2–3 wk and the plants were produced 3–4 wk after anther plating. In the improved system, as much as 53% of the anthers of Pavon 76 were responsive with multiple embryos. For plant regeneration, as many as 22 green and 25 albino plants were produced from 100 anthers. Sixty-five green plants were grown to maturity and 32 (49%) plants were fertile and produced seeds (indicating spontaneous chromosome doubling) while 33 plants did not produce seed. Of five Nebraska breeding lines tested using the protocol, NE96675 was very responsive and the other lines less so, indicating that the protocol is genotype-dependent.