Flame Seedless葡萄胚珠、胚乳及胚发育与败育的研究

对Flame　Seedless葡萄胚珠、胚乳及胚的发育和败育过程进行了系统的细胞学观察,并以有核葡萄品种北醇为对照对Flame　Seedless的胚珠重量及纵横径进行了测定。结果表明：（1）Flame　Seedless内、外珠被及珠孔发育异常,20％～30％的子房未出现受精现象,受精的胚珠重量和纵横径随着胚珠的发育,先呈增加趋势,在某一时期达到最高值,随后下降,最终退化为痕迹。（2）大约有30％的胚乳核能够进行正常分裂,形成胚乳组织,胚乳在花后30d开始退化,其余胚乳核分裂异常。合子在盛花后第19天进行第一次分裂,经过二细胞原胚→多细胞原胚→球形胍,此后胚开始退化;也有的合子始终不发生分裂,最终退化。（3）由于胚乳分裂异常和提早退化,胚缺乏营养致使其发育中止和败育。（4）Flame　Seedless　胚挽救的最佳接种时期在花后第37天。     

无核葡萄的育种与胚挽救技术的运用

植物的胚挽救技术在无核葡萄的育种研究中发展迅速。利用假单性结实的无核葡萄作母本,无核品种作父本,杂交后在合子胚未败育时,进行胚挽救即可获得杂种实生苗,无核率可达82％．理论上可达100％。还介绍了运用该技术时杂交亲本的选择、胚挽救最佳胚龄的确定及相应的培养基成分、激素等技术方法。     

葡萄胚胎发育与败育过程中胚珠的多胺含量变化

花后40~60d期间,种子败育型无核葡萄胚珠的3种多胺含量明显低于有核葡萄,据此认为无核葡萄胚珠中多胺含量急剧下降和较低的多胺水平可能是导致葡萄胚胎败育的重要原因。(Spd Spm)/Put和Spm/PAs比值变化与葡萄胚胎发育密切相关,较低的比值不利于胚胎的正常分化。     

无核香味葡萄胚挽救育种研究

为创制无核香味葡萄新种质,该研究以7个无核葡萄品种为母本,4个玫瑰香味品种为父本,配置了9个杂交组合。田间杂交后,通过胚挽救技术离体培养杂种胚珠,使胚珠中幼胚继续发育,继而在幼胚萌发成苗后利用无核标记对F1代株系进行无核性状的早期检测,同时对杂种幼苗进行温室炼苗,最终移栽大田。结果表明:(1)利用胚挽救技术获得365个胚挽救株系,移栽大田并成活182株杂种苗。(2)对其中72个株系利用无核基因探针GSLP1-569与无核分子标记SCF27-2000进行无核性状检测,分别检测出8个和38个杂种后代携带无核特异性条带,初步确定为无核株系。(3)杂交组合中以‘红宝石无核’、‘火焰无核’、‘昆香无核’为母本的组合成苗率相对较高,较适宜做母本;香味品种中‘玫瑰香’更适合做为父本选育香味无核种质。(4)以‘火焰无核’和‘昆香无核’为母本的杂交组合分别在授粉后的43d和51d采样,胚发育率(6.96%~8.54%和16.92%~18.13%)和成苗率(6.45%和8.75%~10%)较高;‘红宝石无核’ב玫瑰香’、‘莫莉莎无核’ב玫瑰香’和‘波尔莱特’ב玫瑰香’3个组合分别在授粉后59~60d、54d、52d采样,胚挽救育种效果较好;杂交组合‘克瑞森无核’ב巨玫瑰’和‘奇妙无核’ב昆香无核’胚败育时期分别为授粉后39d和48d。(5)幼胚发育至子叶型胚时最易成苗,延长胚珠离体暗培养时间显著增加了畸形胚比率。研究认为,葡萄胚挽救育种过程中,胚珠离体培养时间在8~10周为宜。     

葡萄胚珠,胚乳及胚的发育

本文研究了“早玫瑰”和“新玫瑰”葡萄胚珠、胚乳和胚的发育。结果表明:花后3天胚珠即开始迅速生长,其生长的最终大小依品种成熟期的不同而各异。胚乳游离核在花后33天成为细胞状态。受精后16—21天,合子才开始第一次分裂。当胚乳充满珠心组织时,胚开始迅速发育并一直持续到果实成熟.胚的发育与果实的发育无明显竞争关系。     

葡萄胚珠、胚乳及胚的发育

本文研究了“早玫瑰”和“新玫瑰”葡萄胚珠、胚乳和胚的发育。结果表明：花后3天胚珠即开始迅速生长,共生长的最终大小依品种成熟期的不同而各异。 胚乳游离核在花后33天成为细胞状态。受精后16—21天,合子才开始第一次分裂。当胚乳充满珠心组织时,胚开始迅速发育并—直持续到果实成熟。胚的发育与果实的发育无明显竞争关系。     

无核葡萄胚挽救影响因素的研究

分别以‘底莱特’和‘大粒红无核’自交、‘红宝石无核’ב新郁’和‘爱莫无核’ב黑奥林’胚珠为材料进行无核葡萄胚挽救影响因素的研究。结果显示:(1)‘底莱特’自交和‘红宝石无核’ב新郁’胚珠剥胚后萌发率分别为11.4%和10.0%,高于横切胚珠情况下的萌发率(5.5%、0.6%);(2)用Nitsch培养基培养的‘大粒红无核’自交和‘爱莫无核’ב黑奥林’胚珠的胚萌发率分别为9.5%和5.6%,胚状体诱导率分别为6.3%和3.9%,明显高于ER培养基的萌发率。结果表明:剥胚有利于无核葡萄胚的萌发;Nitsch比ER培养基更有利于胚萌发和胚状体诱导;同时,由于杂交亲本染色体倍性的差异导致胚萌发率不同,表现为:二倍体无核自交>二倍体无核×二倍体有核>二倍体无核×四倍体有核。     

焦核与大核龙眼雌配子体发育、授粉受精及早期胚胎发育的比较研究

利用人工授粉,采用压片法对大核龙眼‘九月乌’和焦核龙眼‘闽焦64-1’、‘闽焦64-2’、‘白核’等的自交与杂交后花粉管的生长特性进行研究,同时应用常规石蜡切片技术对大核与焦核龙眼的雌配子体以及合子胚早期发育进行观察。结果表明,龙眼胚珠在单核胚囊形成前就开始败育,且焦核品种(系)的败育率显著高于大核品种。不同亲本组合的授粉率存在差异,所有授粉组合在授粉36～48 h后均能观察到1个花粉管生长并进入胚囊受精。焦核品种(系)的胚胎在谢花后10 d开始败育,且败育率明显高于大核品种。受精是龙眼子房发育的首要条件,胚珠败育的雌蕊在谢花后10 d不膨大,不能发育形成焦核果实。谢花后10～30 d的早期胚胎败育是形成焦核龙眼的主要原因。焦核品种‘白核’胚乳具有成胚能力。约有24%的‘闽焦64-1’胚珠在胚胎发育过程中,其助细胞、合点端细胞及胚乳发生异常,这可能与早期胚胎败育有关。     

离体培养柑桔小胚获得三倍体植株

三倍体植株因不能进行正常的减数分裂,常产生败育的胚囊和花粉;又因柑桔属植物具有单性结实的能力,因而人工培育三倍体植株已成为获得无核柑桔品种的一条重要途径。在柑桔属植物中,获得三倍体主要有三条途径:(1)通过胚乳离休培养诱导三倍体植株;(2)四倍体品种同二倍体品种杂交,获得三倍体;     

离体培养柑桔小胚获得三倍体植株

兰倍休植株因不能进行正常的减数分裂, 常产生败育的胚囊和花粉;又因柑桔属植物具 有单性结实的能力,因而人工培育三倍体植株 已成为获得无核柑桔品种的一条重要途径。在 柑桔属植物中,获得三倍体主要有三条途径： (1)通过胚乳离体培养诱导三倍体植株［[I>; (2) 四倍体品种同二倍体品种杂交,获得三倍体[41 . (3）单胚的二倍体柑桔品种作母本进行杂交或 自然授粉后,可产生一定比例的,重量为正常种 子1/6至123的小粒种子,并已确定此小粒种 子是三倍体［[2,3,8,10]。这无疑为柑桔的三倍体培育 提供了一条简便、实用和快速的新途径。     

The metacaspase gene family of Vitis vinifera L.: Characterization and differential expression during ovule abortion in stenospermocarpic seedless grapes

In both plants and animals, programmed cell death (PCD) is an indispensable process that removes redundant cells. In seedless grapes (Vitis vinifera), abnormal PCD in ovule cells and subsequent ovule abortion play key roles in stenospermocarpy. Metacaspase, a type of cysteine-dependent protease, plays an essential role in PCD. To reveal the characteristics of the metacaspase (MC) gene family and the relationship between metacaspases and the seedless trait, we identified the 6 V. vinifera metacaspases VvMC1–VvMC6, from the grape genome, using BLASTN against the 9 known Arabidopsis metacaspases. We also obtained full-length cDNAs by RT-PCR. Each of the 6 grape metacaspases contains small (p10-like) and a large (p20-like) conserved structural domains. Phylogenetic analysis of 6 grape and 9 Arabidopsis metacaspases showed that all metacaspases could be grouped into two classes: Type I and Type II. Each phylogenetic branch shares a similar exon/intron structure. Furthermore, the putative promoters of the grape metacaspases contained cis-elements that are involved in grape endosperm development. Moreover, expression analysis of metacaspases using real-time quantitative PCR demonstrated that VvMC1 and VvMC2 were able to be detected in any tissue, and VvMC3, VvMC4, VvMC5 and VvMC6 exhibited tissue-specific expression. Lastly, in cv. Thompson seedless grapes VvMC1, VvMC3, and VvMC4 were significantly up-regulated at the 35 DAF during ovule development, roughly same stage as endosperm abortion. In addition, the expression trend of VvMC2 and VvMC5 was similar between cv. Pinot Noir and cv. Thompson grape ovule development and that of VvMC6 was sustained in a relatively low level except the expression of cv. Pinot Noir significantly up-regulated in 25 DAF. Our data provided new insights into PCD by identifying the grape metacaspase gene family and provide a useful reference for further functional analysis of metacaspases in grape.     

DEVELOPMENT OF NORMAL AND SEEDLESS ACHENES IN CICHORIUM INTYBUS (COMPOSITAE)

Cross- and partially cross-pollinated capitula of Cichorium intybus (Compositae, Lactuceae) were examined for a study of normal and seedless fruit development respectively. Embryos develop according to the Asterad pattern, and the free-nuclear endosperm becomes cellular 15–17 hrs after pollination. A zone of disorganized cellular material surrounds the embryo sac at anthesis, and, in normal achenes, this zone expands as the seed develops. Initially the developing seed elongates and comes into contact with the top of the ovary by 48 hrs. In contrast to this pattern, the ovule in developing seedless achenes degenerates within 72 hrs. Irregularities, such as an abnormally proliferating endothelium, embryo formation without endosperm, and endosperm formation without an embryo often accompany this degeneration. Differentiation of the pericarp in seeded achenes begins between 48 and 72 hrs, starting at the apex and proceeding basipetally; in seedless fruits the process is similar though initiated somewhat later. The normal pericarp at maturity exhibits a pigmented exocarp, a broad mesocarp of thick-walled lignified cells, and a tenuous endocarp. In seedless achenes the fruit coat is similar except that the exocarp is colorless and the cells of the mesocarp are relatively small.     

SEED DEVELOPMENT IN CITRUS WITH SPECIAL REFERENCE TO 2X X 4X CROSSES

Comparative cytological and histological studies during embryogenesis of seeds from 2x X 2x and 2x x 4x crosses indicate that the ratio of ploidy level between embryo and endosperm is the most important factor affecting the course of seed development. The crosses produced seeds with the expected ploidy relationships between embryo, endosperm, and maternal tissue of 2:3:2 and 3:4:2 as well as the anomalous relationships 3:5:2, 4:6:2, and 6:10:2. All but 3:4:2 resulted in normal, germinable seeds. The ploidy level of the maternal tissue in relation to that of the embryo or endosperm did not appear to have any effect on seed development. About 92–99 % of seeds from 2x x 4x crosses containing triploid embryos with tetraploid endosperm aborted at different stages of embryogenesis. The abortion in all cases was preceded by abnormalities in the tetraploid endosperm. It is postulated that the unbalance of chromosome number between embryo and endosperm disturbs physiological relationships between these two tissues, leading first to the abortion of the endosperm and then of the embryo.     

Imprinting of the MEDEA polycomb gene in the Arabidopsis endosperm.

In flowering plants, two cells are fertilized in the haploid female gametophyte. Egg and sperm nuclei fuse to form the embryo. A second sperm nucleus fuses with the central cell nucleus that replicates to generate the endosperm, which is a tissue that supports embryo development. MEDEA (MEA) encodes an Arabidopsis SET domain Polycomb protein. Inheritance of a maternal loss-of-function mea allele results in embryo abortion and prolonged endosperm production, irrespective of the genotype of the paternal allele. Thus, only the maternal wild-type MEA allele is required for proper embryo and endosperm development. To understand the molecular mechanism responsible for the parent-of-origin effects of mea mutations on seed development, we compared the expression of maternal and paternal MEA alleles in the progeny of crosses between two Arabidopsis ecotypes. Only the maternal MEA mRNA was detected in the endosperm from seeds at the torpedo stage and later. By contrast, expression of both maternal and paternal MEA alleles was observed in the embryo from seeds at the torpedo stage and later, in seedling, leaf, stem, and root. Thus, MEA is an imprinted gene that displays parent-of-origin-dependent monoallelic expression specifically in the endosperm. These results suggest that the embryo abortion observed in mutant mea seeds is due, at least in part, to a defect in endosperm function. Silencing of the paternal MEA allele in the endosperm and the phenotype of mutant mea seeds supports the parental conflict theory for the evolution of imprinting in plants and mammals.     

四倍体大燕麦与六倍体裸燕麦的受精作用和胚胎发育

用石蜡切片法,对四倍体大燕麦（Avena magna L.）和六倍体裸燕麦（Avena nuda L.）杂交的受精作用和胚胎发育进行了观察。结果表明,六倍体裸燕麦花偻在四倍体大燕麦柱头上萌发良好,花粉管可顺利长入花柱和胚囊,观察的168个四倍体大燕麦子房中,2.38%发生了双受精,产生了胚和胚乳;1.79%发生了单卵受精,只产生胚乳而无胚;总受精率为5.36%;成胚率为4.17%。由于胚乳的缺管或发育异常及败育,最终难以获得有生活力的种子,为四倍体大燕麦和六倍体裸燕麦杂交提供了细胞胚胎学证据。     

无核葡萄品种的无核性来源分析

系谱分析表明,无核白是世界范围内无核葡萄育种的主要无核性来源,以其为最早的亲本材料,经过简单杂交或多亲多代杂交,培育了一系列的无核葡萄品种;无核紫和俄罗斯无核是部分无核品种的原始亲本材料之一;有核葡萄的无核芽变也是无核品种选育的重要途径。     

Embryology of Early Abortion Due to Limited Maternal Resources in Pisum sativum L.

In most flowering plants, many embryos are aborted early intheir development due to limited maternal resources. The kin-conflictinterpretation of plant embryology predicts these abortionsshould be under maternal control. In a study of the abortionprocess in Pisum sativum, we found the first visible indicationof abortion was formation of a weak hypostase. Callose was locallydeposited around the chalazal endosperm haustorium, and ligninalong the outer cell walls of the remnant nucellar tissue. Thenucellus was compressed by proliferating adjacent inner integumentalcells. The endosperm haustorium's cytoplasm was forced backinto the embryo sac cavity. With suppression of haustorial activitythe endosperm nuclei gradually enlarged followed by enlargementof the embryo and suspensor nuclei. Finally, nuclei and cytoplasm throughout the endosperm and embryolost stainability and broke down. Four successive stages wererecognized in seed abortion. In seeds developing to maturity,no hypostase was developed and the haustorium continued to digestboth the remnant nucellus and the proliferated inner integumentalcells. These observations are consistent with the kin-conflicthypothesis. Pisum sativum, garden pea, ovule abortion, histology, hypostase, kin-conflict hypothesis     

四倍体大燕麦与六倍体裸燕麦的受精作用和胚胎发育

用石蜡切片法,对四倍体大燕麦(Avena magna L.)和六倍体裸燕麦(Avena nuda L.)杂交的受精作用和胚胎发育进行了观察。结果表明,六倍体裸燕麦花粉在四倍体大燕麦柱头上萌发良好,花粉管可顺利长入花柱和胚囊。观察的168个四倍体大燕麦子房中,238%发生了双受精,产生了胚和胚乳;179%发生了单卵受精,只产生胚乳而无胚;总受精率为536%;成胚率为417%。由于胚乳的缺乏或发育异常及败育,最终难以获得有生活力的种子。为四倍体大燕麦和六倍体裸燕麦杂交提供了细胞胚胎学证据。