,*中国科学院植物研究所, 植物分子生理学重点实验室, 北京 100093Chinese Scientists Make Groundbreaking Discoveries in Clonal Propagation of F1 Hybrids
Zhihui Xue, Kang Chong,*Key Laboratory of Plant Molecular Physiology, Institute of Botany, Chinese Academy of Sciences, Beijing 100093, China通讯作者:
收稿日期:2019-01-9接受日期:2019-01-10网络出版日期:2019-01-30
Corresponding authors:
Received:2019-01-9Accepted:2019-01-10Online:2019-01-30
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薛治慧, 种康. 中国科学家在杂种F1克隆繁殖研究领域取得突破性进展. 植物学报, 2019, 54(1): 1-3 doi:10.11983/CBB19004
Xue Zhihui, Chong Kang.
杂种优势(heterosis)是指遗传基础不同的亲本, 其杂交子一代在生物量、产量和抗性等方面优于双亲的遗传现象。自20世纪发现杂种优势以来, 已在玉米(Zea mays)、高粱(Sorghum vulgare)、水稻(Oryza sativa)、油菜(Brassica campestris)及向日葵(Helianthus annuus)等主要农作物中得到广泛应用, 对于提高世界范围内粮食单产发挥了重要作用, 为解决粮食安全问题做出了重要贡献。
杂交稻是杂种优势利用最为成功的作物之一, 自20世纪70年代成功研制以来, 已经历一系列质的飞跃。第一代杂交稻是袁隆平等以质核互作型雄性不育系(cytoplasmic male sterility)为遗传工具, 创制的“三系法”杂交水稻, 其单产比常规水稻品种增产20%左右, 是水稻育种历史上的一次重要突破, 为保障我国粮食安全发挥了重要作用。第二代杂交稻是以光温敏雄性不育系(photothermo-sensitive male sterility)为遗传材料, 创制的“两系法”杂交水稻。由于“两系法”杂交稻不存在不育基因与恢复基因间恢保关系的限制, 育成优良组合的概率要远高于“三系法”杂交稻。“一系法”则是通过无融合生殖(apomixis)来固定杂种优势, 理论上是水稻杂种优势利用的最佳途径(Yuan, 2018)。
由于无融合生殖没有经过受精过程, 直接产生种子, 因而其后代与亲本基因型完全一致。目前, 已报道有400多种被子植物(angiosperm)具有无融合生殖现象, 但是在主要农作物中尚未发现有无融合生殖现象(Koltunow and Grossniklaus, 2003)。科学家通过对自然界中存在无融合生殖的物种进行遗传分析, 获得了无融合生殖现象的遗传模式及其调控位点(Ozias- Akins and van Dijk, 2007; Barcaccia and Albertini, 2013)。然而, 研究者曾尝试将调控无融合生殖的染色体片段导入相应的栽培系中, 但并未获得成功(Barcaccia and Albertini, 2013)。此外, 研究者也尝试通过对控制有性生殖(sexual reproduction)过程的基因进行突变, 从头创制无融合生殖生物(Spillane et al., 2004)。
减数分裂(meiosis)是真核生物(eukaryote)有性生殖过程中发生的核心生物学事件, 对于有性生殖过程的正常进行有着极其重要的意义。在减数分裂过程中, 同源染色体的非姊妹染色单体间发生交换, 是杂交后代遗传多样性形成的生物学基础。Raphae?l Mercier研究组分别在拟南芥(Arabidopsis thaliana)和水稻中同时突变3个调控减数分裂过程的基因, 获得了名为MiMe (Mitosis instead of Meiosis)的突变体。MiMe孢母细胞的减数分裂过程被类似的有丝分裂(mitosis)过程所取代, 不发生同源染色体交换, 因而产生克隆二倍体(diploid)的雄性与雌性配子(Marimuthu et al., 2011; Mieulet et al., 2016)。由于MiMe植株的自交后代染色体自发加倍, 进一步将拟南芥MiMe植株和CenH3-介导的染色体消除系进行杂交, 成功获得克隆的二倍体后代。
中国水稻研究所王克剑研究组通过多重CRISPR- Cas9基因编辑技术, 对杂交稻组合春优84中PAIR1、REC8和OSD1这3个减数分裂相关基因进行编辑, 成功获得了MiMe材料。MiMe在性细胞形成过程中不发生同源染色体重组(homologous recombination), 产生与亲本基因型一致的二倍体配子(Wang et al., 2019)。已有研究表明, 玉米MTL (MATRILINEAL)基因可以诱导产生单倍体(haploid)植株(Gilles et al., 2017; Kelliher et al., 2017; Liu et al., 2017)。与拟南芥中通过CenH3诱导孤雌生殖(parthenogenesis)相比, 其优点是mtl突变体植株自交可直接获得孤雌生殖单倍体后代。水稻中存在玉米MTL的同源基因, 且其生物学功能与玉米MTL相同(Yao et al., 2018)。王克剑研究组还通过CRISPR-Cas9系统验证了该基因的功能, 并成功诱导出单倍体植株。在此基础上, 他们同时对春优84的PAIR1、REC8、OSD1和MTL基因进行编辑, 成功获得具有无融合生殖现象的Fix (Fixation of hybrids)材料(图1), 并获得与杂交亲本基因型一致的克隆种子。对其二倍体后代进行基因组重测序分析表明, 克隆繁殖的二倍体植株与杂交种春优84基因型一致, 说明杂合背景的基因型能在不同有性世代间传递。
图1
新窗口打开|下载原图ZIP|生成PPT图1利用基因编辑获得克隆杂交种的模式图(改自Wang et al., 2019)
Figure 1A model for fixation of heterozygosity of the hybrid by genome editing (modified from Wang et al., 2019)
常规有性生殖过程(上图)中, 通过重组的单倍体配子融合产生重组的二倍体后代。克隆繁殖策略(下图)基于两点: 一是通过基因编辑获得的MiMe材料可以产生克隆二倍体配子; 二是通过敲除MTL基因使性细胞进行单性生殖。克隆植株(Fix)产生的克隆杂交种与杂交亲本基因型一致。
In natural sexual reproduction (up), recombined diploid progeny are produced by fusion of recombined haploid game-tes. The clonal reproduction strategy (bottom) is based on two events: MiMe leads to the formation of clonal diploid gametes, and parthenogenesis of the diploid by knocking out the MTL gene. The clonal plant (Fix) produces clonal progeny that are genetically identical to the hybrid parent.
Wang等(2019)的研究工作证实了通过无融合生殖固定水稻杂种优势的可能性, 为实现“一系法”杂种优势利用探索出一种可能的技术路径。未来通过对该技术的进一步完善与推广, 可望极大地减少农作物育种的工作量, 缩短良种培育周期, 为农作物杂种优势的高效利用带来光明前景。
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被引期刊影响因子
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DOI:10.15252/embj.201796603URLPMID:28228439 [本文引用: 1]
Gynogenesis is an asexual mode of reproduction common to animals and plants, in which stimuli from the sperm cell trigger the development of the unfertilized egg cell into a haploid embryo. Fine mapping restricted a major maize QTL (quantitative trait locus) responsible for the aptitude of inducer lines to trigger gynogenesis to a zone containing a single gene NOT LIKE DAD (NLD) coding for a patatin-like phospholipase A. In all surveyed inducer lines, NLD carries a 4-bp insertion leading to a predicted truncated protein. This frameshift mutation is responsible for haploid induction because complementation with wild-type NLD abolishes the haploid induction capacity. Activity of the NLD promoter is restricted to mature pollen and pollen tube. The translational NLD::citrine fusion protein likely localizes to the sperm cell plasma membrane. In Arabidopsis roots, the truncated protein is no longer localized to the plasma membrane, contrary to the wild-type NLD protein. In conclusion, an intact pollen-specific phospholipase is required for successful sexual reproduction and its targeted disruption may allow establishing powerful haploid breeding tools in numerous crops.
DOI:10.1038/nature20827URLPMID:28114299 [本文引用: 1]
Abstract Sexual reproduction in flowering plants involves double fertilization, the union of two sperm from pollen with two sex cells in the female embryo sac. Modern plant breeders increasingly seek to circumvent this process to produce doubled haploid individuals, which derive from the chromosome-doubled cells of the haploid gametophyte. Doubled haploid production fixes recombinant haploid genomes in inbred lines, shaving years off the breeding process. Costly, genotype-dependent tissue culture methods are used in many crops, while seed-based in vivo doubled haploid systems are rare in nature and difficult to manage in breeding programmes. The multi-billion-dollar maize hybrid seed business, however, is supported by industrial doubled haploid pipelines using intraspecific crosses to in vivo haploid inducer males derived from Stock 6, first reported in 1959 (ref. 5), followed by colchicine treatment. Despite decades of use, the mode of action remains controversial. Here we establish, through fine mapping, genome sequencing, genetic complementation, and gene editing, that haploid induction in maize (Zea mays) is triggered by a frame-shift mutation in MATRILINEAL (MTL), a pollen-specific phospholipase, and that novel edits in MTL lead to a 6.7% haploid induction rate (the percentage of haploid progeny versus total progeny). Wild-type MTL protein localizes exclusively to sperm cytoplasm, and pollen RNA-sequence profiling identifies a suite of pollen-specific genes overexpressed during haploid induction, some of which may mediate the formation of haploid seed. These findings highlight the importance of male gamete cytoplasmic components to reproductive success and male genome transmittance. Given the conservation of MTL in the cereals, this discovery may enable development of in vivo haploid induction systems to accelerate breeding in crop plants.
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DOI:10.1126/science.1199682URLPMID:21330535 [本文引用: 1]
Abstract Cloning through seeds has potential revolutionary applications in agriculture, because it would allow vigorous hybrids to be propagated indefinitely. However, asexual seed formation or apomixis, avoiding meiosis and fertilization, is not found in the major food crops. To develop de novo synthesis of apomixis, we crossed Arabidopsis MiMe and dyad mutants that produce diploid clonal gametes to a strain whose chromosomes are engineered to be eliminated after fertilization. Up to 34% of the progeny were clones of their parent, demonstrating the conversion of clonal female or male gametes into seeds. We also show that first-generation cloned plants can be cloned again. Clonal reproduction through seeds can therefore be achieved in a sexual plant by manipulating two to four conserved genes.
DOI:10.1038/cr.2016.117URLPMID:5099866
Introduction of clonal reproduction through seeds (apomixis) in crops has the potential to revolutionize agriculture by allowing self-propagation of any elite variety, in particular F1 hybrids. In the sexual model plant Arabidopsis thaliana synthetic clonal reproduction through seeds can be artificially implemented by (i) combining three mutations to turn meiosis into mitosis (MiMe) and (ii) crossing the obtained clonal gametes with a line expressing modified CENH3 and whose genome is eliminated in the zygote. Here we show that additional combinations of mutations can turn Arabidopsis meiosis into mitosis and that a combination of three mutations in rice (Oryza sativa) efficiently turns meiosis into mitosis, leading to the production of male and female clonal diploid gametes in this major crop. Successful implementation of the MiMe technology in the phylogenetically distant eudicot Arabidopsis and monocot rice opens doors for its application to any flowering plant and paves the way for introducing apomixis in crop species.
DOI:10.1146/annurev.genet.40.110405.090511URLPMID:18076331 [本文引用: 1]
Abstract Apomixis, asexual reproduction through seeds, has the potential to revolutionize agriculture if its genetic basis can be elucidated. However, the genetic control of natural apomixis has remained obscure until quite recently, owing to all the complications of Mendelian genetics, such as epistatic gene interactions, components that are expressed sporophytically and gametophytically, expression modifiers, polyploidy, aneuploidy, segregation distortion, suppressed recombination, etc., that seem to have accumulated during the evolution of apomixis. In this review we show how molecular markers and superior phenotypic methods have been used to clarify the genetics of apomixis in monocots as well as dicots during the past 15 years. Many of the complexities in the genetics of apomixis are likely secondary and have evolved as a consequence of the reproductive process. New mapping techniques, such as comparative mapping, linkage disequilibrium mapping, and deletion mapping, and new high-throughput sequencing methods, will help to penetrate the core of apomixis chromosomal regions. If the evolutionary genetic load can be exposed and removed, the apomixis genes may be used in agriculture as a tool to fix elite genotypes.
DOI:10.1038/nbt976URLPMID:15175691 [本文引用: 1]
Apomixis is the process of through seed, in the absence of meiosis and , generating clonal progeny of maternal origin. Major benefits to agriculture could result from harnessing apomixis in crop . Although >400 apomictic plant species are known, apomixis is rare among crop , and the transfer of apomixis to crop varieties by conventional breeding has been largely unsuccessful. Because apomictic and sexual pathways are closely related, de novo engineering of apomixis might be achieved in sexually reproducing crops. Early consideration of issues relating to biosafety and intellectual property (IP) management can facilitate the acceptance and deployment of apomixis technology in agriculture.
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Apomixis in plant repro- duction: a novel perspective on an old dilemma
2
2013
... 由于无融合生殖没有经过受精过程, 直接产生种子, 因而其后代与亲本基因型完全一致.目前, 已报道有400多种被子植物(angiosperm)具有无融合生殖现象, 但是在主要农作物中尚未发现有无融合生殖现象(
... ).然而, 研究者曾尝试将调控无融合生殖的染色体片段导入相应的栽培系中, 但并未获得成功(
Loss of pollen-specific phospholipase NOT LIKE DAD triggers gynogenesis in maize
1
2017
... 中国水稻研究所王克剑研究组通过多重CRISPR- Cas9基因编辑技术, 对杂交稻组合春优84中PAIR1、REC8和OSD1这3个减数分裂相关基因进行编辑, 成功获得了MiMe材料.MiMe在性细胞形成过程中不发生同源染色体重组(homologous recombination), 产生与亲本基因型一致的二倍体配子(
MATRILINEAL, a sperm-specific phospholipase, triggers maize haploid in- duction
1
2017
... 中国水稻研究所王克剑研究组通过多重CRISPR- Cas9基因编辑技术, 对杂交稻组合春优84中PAIR1、REC8和OSD1这3个减数分裂相关基因进行编辑, 成功获得了MiMe材料.MiMe在性细胞形成过程中不发生同源染色体重组(homologous recombination), 产生与亲本基因型一致的二倍体配子(
Apomixis: a develop-mental perspective
1
2003
... 由于无融合生殖没有经过受精过程, 直接产生种子, 因而其后代与亲本基因型完全一致.目前, 已报道有400多种被子植物(angiosperm)具有无融合生殖现象, 但是在主要农作物中尚未发现有无融合生殖现象(
A 4 bp insertion at ZmPLA1 encoding a putative phospholipase A generates haploid induction in maize.
2
2017
... 中国水稻研究所王克剑研究组通过多重CRISPR- Cas9基因编辑技术, 对杂交稻组合春优84中PAIR1、REC8和OSD1这3个减数分裂相关基因进行编辑, 成功获得了MiMe材料.MiMe在性细胞形成过程中不发生同源染色体重组(homologous recombination), 产生与亲本基因型一致的二倍体配子(
... A model for fixation of heterozygosity of the hybrid by genome editing (
Synthetic clonal reproduction through seeds
1
2011
... 减数分裂(meiosis)是真核生物(eukaryote)有性生殖过程中发生的核心生物学事件, 对于有性生殖过程的正常进行有着极其重要的意义.在减数分裂过程中, 同源染色体的非姊妹染色单体间发生交换, 是杂交后代遗传多样性形成的生物学基础.Raphae?l Mercier研究组分别在拟南芥(Arabidopsis thaliana)和水稻中同时突变3个调控减数分裂过程的基因, 获得了名为MiMe (Mitosis instead of Meiosis)的突变体.MiMe孢母细胞的减数分裂过程被类似的有丝分裂(mitosis)过程所取代, 不发生同源染色体交换, 因而产生克隆二倍体(diploid)的雄性与雌性配子(
Turning rice meiosis into mitosis
0
2016
Mendelian genetics of apomixis in plants
1
2007
... 减数分裂(meiosis)是真核生物(eukaryote)有性生殖过程中发生的核心生物学事件, 对于有性生殖过程的正常进行有着极其重要的意义.在减数分裂过程中, 同源染色体的非姊妹染色单体间发生交换, 是杂交后代遗传多样性形成的生物学基础.Raphae?l Mercier研究组分别在拟南芥(Arabidopsis thaliana)和水稻中同时突变3个调控减数分裂过程的基因, 获得了名为MiMe (Mitosis instead of Meiosis)的突变体.MiMe孢母细胞的减数分裂过程被类似的有丝分裂(mitosis)过程所取代, 不发生同源染色体交换, 因而产生克隆二倍体(diploid)的雄性与雌性配子(
Apomixis technology development-virgin births in farmers' fields?
1
2004
... 由于无融合生殖没有经过受精过程, 直接产生种子, 因而其后代与亲本基因型完全一致.目前, 已报道有400多种被子植物(angiosperm)具有无融合生殖现象, 但是在主要农作物中尚未发现有无融合生殖现象(
Clonal seeds from hybrid rice by simultaneous genome engineering of meiosis and fertilization genes
1
... 由于无融合生殖没有经过受精过程, 直接产生种子, 因而其后代与亲本基因型完全一致.目前, 已报道有400多种被子植物(angiosperm)具有无融合生殖现象, 但是在主要农作物中尚未发现有无融合生殖现象(
OsMATL mutation induces haploid seed formation in indica rice
1
2018
... 中国水稻研究所王克剑研究组通过多重CRISPR- Cas9基因编辑技术, 对杂交稻组合春优84中PAIR1、REC8和OSD1这3个减数分裂相关基因进行编辑, 成功获得了MiMe材料.MiMe在性细胞形成过程中不发生同源染色体重组(homologous recombination), 产生与亲本基因型一致的二倍体配子(
The strategy for hybrid rice development
4
2018
... 杂交稻是杂种优势利用最为成功的作物之一, 自20世纪70年代成功研制以来, 已经历一系列质的飞跃.第一代杂交稻是袁隆平等以质核互作型雄性不育系(cytoplasmic male sterility)为遗传工具, 创制的“三系法”杂交水稻, 其单产比常规水稻品种增产20%左右, 是水稻育种历史上的一次重要突破, 为保障我国粮食安全发挥了重要作用.第二代杂交稻是以光温敏雄性不育系(photothermo-sensitive male sterility)为遗传材料, 创制的“两系法”杂交水稻.由于“两系法”杂交稻不存在不育基因与恢复基因间恢保关系的限制, 育成优良组合的概率要远高于“三系法”杂交稻.“一系法”则是通过无融合生殖(apomixis)来固定杂种优势, 理论上是水稻杂种优势利用的最佳途径(
... 中国水稻研究所王克剑研究组通过多重CRISPR- Cas9基因编辑技术, 对杂交稻组合春优84中PAIR1、REC8和OSD1这3个减数分裂相关基因进行编辑, 成功获得了MiMe材料.MiMe在性细胞形成过程中不发生同源染色体重组(homologous recombination), 产生与亲本基因型一致的二倍体配子(
... 利用基因编辑获得克隆杂交种的模式图(
...
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