Analysis of genetic diversity and discrimination of Oil Palm DxP populations based on the origins of pisifera elite parents

A total of 251 Dura cross Pisifera (DxP) hybrid palms from six populations descending from six parental African Pisifera origins and involving 12 progenies were analyzed with 19 selected Simple Sequence Repeats (SSR) markers. A total of 110 alleles were produced, ranging from three to eight per SSR, with a mean of 5.8 alleles per SSR locus. Of these, 68.5% were considered shared alleles by more than one population and the remaining 31.5% were population specific alleles. They generated between six and 21 haplotypes in all populations, and depending on the SSR marker, between one and 10 haplotypes within populations. Various parameters for analyzing genetic variability, differentiation and genetic structure were computed using GenAlEx, Structure and Darwin software. The obtained results confirmed microsatellites as a robust, feasible and trustful method for obtaining DNA fingerprints, tracing the source of oil palm samples. With respect to the authenticity of materials or for solving legitimacy issues, accession belonging to each population by SSR markers could be distinguished, but additional SSR should be screened for improving this process.     

结球甘蓝花粉类钙调素蛋白基因BoCML49的克隆及表达分析

以结球甘蓝E1为材料,提取花粉萌发前和萌发后的混合花粉总蛋白,总蛋白双向电泳后通过MALDI-TOF-MS鉴定分析差异点,根据差异点分析结果,利用同源克隆得到甘蓝BoCML49基因707 bp的cDNA片段。通过对BoCML49基因的qPCR分析表明其表达量在花粉萌发前约为萌发后的2.73倍,表明BoCML49基因在花粉萌发后表达下调。通过5¢-Race和3¢-Race分别得到550 bp和721 bp大小cDNA片段,最终得到结球甘蓝BoCML49全长cDNA序列,开放阅读框位于125~
1 078 bp处,加尾信号(AATAAA)位于第1 222 bp处。通过cDNA推导得到的氨基酸序列分析表明,BoCML49编码317个氨基酸残基,预测分子量为33.51 kD,pI为6.93,经Smart-embl预测其含2个重要的EF-hand结构,分别位于序列第150~178和第216~244位氨基酸残基处,预测结果显示其含有7个α-螺旋和10个β-折叠结构。系统发育树表明结球甘蓝BoCML49与拟南芥AtCML49和AtCML50的亲缘关系较近。BoCML49基因的原核表达得到37.55 kD的融合蛋白。     

芒花粉的生活力及测定方法比较

本研究先后采用了花粉离体萌发法、FDA染色法和I2-KI染色法测定了芒离体花粉的生活力。结果表明：在离体萌发法中,离体花粉在培养5 min后即开始萌发,培养30 min后花粉管平均长度达到了145.77 μm。芒离体花粉的平均初始萌发率为82.6 %,但花粉生活力下降很快,室温保存90 min的花粉其萌发率已下降至3.0 %。因此利用离体萌发法能够准确有效地测定芒花粉生活力的变化规律。而利用FDA染色法和I2-KI染色法测定的芒花粉初始生活力与离体萌发法的结果基本一致,分别为84.6 %和86.6 %,但这两种染色法不适合用于跟踪测定花粉生活力的变化。     

Exogenously Applied Plant Growth Regulators Affect Heat‐Stressed Rice Pollens

Increasing temperature due to global warming has emerged one of the gravest threats to rice production. This study examined the influence of high temperature and exogenously applied plant growth regulators on pollen fertility, anther dehiscence, pollen germination and metabolites synthesis in pollens of two rice cultivars (IR‐64 and Huanghuazhan (HHZ)). Plants were subjected to high day temperature (HDT), high night temperature (HNT) and control temperature (CT) in controlled growth chambers. Four different combinations of ascorbic acid (Vc), alpha‐tocopherol (Ve), brassinosteroids (Br), methyl jasmonates (MeJA) and triazoles (Tr) were used along with a nothing applied control. Our results depicted that high temperature severely reduced the pollen fertility, anther dehiscence, pollen retention, germination and metabolites synthesis in pollens of both rice cultivars. Nonetheless, exogenous application of various plant growth regulators assuaged the adverse effects of high temperature and Vc + Ve + MeJA + Br was found the best combination than the other treatments for every studied characteristic. The HNT posed more negative effects than the HDT. Variations were also apparent between cultivars and HHZ performed better than IR‐64 under high‐temperature stress, with higher pollen fertility, better anther dehiscence, and greater pollen retention and germination rates. The greater tolerance of HHZ to high temperature was related with the higher synthesis of metabolites in this cultivar.     

Pollen characteristics and stigma receptivity for <Emphasis Type="Italic">Anemone coronaria</Emphasis> L.

To obtain crossing products, an efficient pollination and subsequent fertilization is essential. This efficient pollination is achieved by pollen germination and tube growth. Here, these pollen characteristics of 2 genetically differentiating cultivars of Anemone coronaria L. were investigated in vitro. For the essential components boron, calcium and an osmoticant, only calcium showed to be crucial for pollen germination. Boron concentrations influenced the pollen tube length with a concentration of 100 mg l^?1 H₃BO₃ resulting in the longest pollen tubes. For the osmoticant sucrose, a concentration of 100 g l^?1 caused a significant positive effect on both pollen germination and pollen tube length for the 2 cultivars. The cultivars reacted similarly with respect to the investigated compounds. Next to this, the pollen development was correlated with 8 different flower stages in A. coronaria. As a final point, the germination of an optimized pollen germination medium was compared with in vivo pollen germination in cross-pollinations within the same cultivar (identified by aniline blue staining). For ‘Mistral Wine’, pollen germination percentage was lower in vitro than in vivo, while ‘Wicabri Blue’ pollen showed no significant difference in germination rates in vivo and in vitro. To achieve fertilization a following requisite is that the stigma is receptive. To study this, the most receptive female flower stage of the 8 different flower stages for A. coronaria was characterized by aniline blue staining. A. coronaria clearly showed protogyny.     

DNA甲基化参与调控马铃薯响应干旱胁迫的关键基因挖掘

植物受到干旱胁迫时,会通过DNA甲基化做出快速反应以帮助其应对胁迫。为探究在干旱胁迫下,DNA甲基化是如何影响基因转录表达,本研究对甘露醇模拟干旱和5-azadC(去甲基化)处理下,抗旱性不同的2个马铃薯品种(抗旱型,青薯9号;干旱敏感型,大西洋)进行转录组学分析,以Fold-change>2和校正后P<0.01进行差异表达基因(DEG)的筛选。GO富集分析发现,2种处理都共同显著富集到氧化应激和碳水化合物代谢过程相关的GO term。说明不同耐旱性马铃薯在响应干旱胁迫时,与这些GO term相关的基因也受DNA去甲基化调控。对既响应干旱又响应DNA去甲基化的1345个DEG进行KEGG功能富集发现,与植物抗旱相关的通路有植物MAPK信号途径、植物激素信号转导途径、植物谷胱甘肽代谢通路、糖酵解与糖异生和磷酸肌醇代谢通路。说明这些通路相关基因在大西洋和青薯9号2个抗旱性不同的马铃薯品种中,响应干旱的敏感性受DNA甲基化调控。接着对DEG上游1500 bp启动子区域进行顺式作用原件和甲基化CpG岛分析发现,干旱胁迫下参与植物谷胱甘肽代谢的GST基因通过DNA去甲基化来降低启动子区ABRE和CAAT-box作用元件的甲基化水平,进而激活该基因的表达以应对干旱胁迫。因此,利用比较转录组学分析干旱和DNA去甲基化处理下的差异基因,可挖掘到DNA甲基化参与调控马铃薯响应干旱胁迫的相关基因,为研究马铃薯干旱胁迫响应的表观遗传学机理提供新的研究思路。     

同源四倍体西瓜自交花粉萌发生长与内源激素的关系

对花粉离体培养和花粉在雌蕊柱头上萌发、生长的研究发现,同源四倍体西瓜自交,花粉萌发率低,花粉管在花柱中生长缓慢,在花粉萌发和生长过程中伴随出现结节状、花粉管尖端膨大呈球形花粉管及双花粉管等异常现象。经过内源激素测定分析表明,上述现象与四倍体西瓜雌芯和花粉中细胞分裂素－－玉米素核苷（ＺＲ）含量过高、雌蕊的ＩＡＡ和ＧＡ３与ＡＢＡ的比值低、及花粉和雌蕊共同作用有关。     

低温胁迫下马铃薯的数字基因表达谱分析

以马铃薯品种合作88为材料,利用数字基因表达谱(DGE)技术,对–2℃低温胁迫处理后的马铃薯叶片c DNA文库进行差异基因表达谱分析。结果表明,有28 505个基因受低温胁迫诱导差异表达,其中上调表达基因13 703个,下调表达基因14 802个。GO功能显著性富集分析表明,DEGs主要涉及信号生物代谢过程、氧化还原过程、能量代谢、次生代谢过程以及催化活性。KEGG富集分析表明,上调表达基因主要富集于苯丙烷、光合作用天线蛋白、类胡萝卜素的生物合成、苯丙氨酸代谢及淀粉与蔗糖代谢途径,而下调表达基因主要富集于植物激素信号转导途径。利用实时荧光定量PCR(q RT-PCR)验证4个DEGs在低温胁迫条件下的差异表达,其结果与DGE分析结果基本一致,证实了DGE测序结果的可靠性。     

植物生长调节剂对‘秦冠’苹果花粉管生长和花粉萌发的影响

研究吲哚乙酸、萘乙酸、赤霉素、2,4-D、6-BA5种植物生长调节剂对苹果花粉管生长和花粉萌发的影响。以贮藏6个月的‘秦冠’花粉为材料,配制不同浓度梯度植物生长调节剂的固体培养基培养花粉,研究不同浓度植物激素对花粉管生长和花粉萌发的影响。结果表明,当吲哚乙酸≤1 mg/L、萘乙酸≤1 mg/L、6-BA≤5 mg/L、赤霉素≤10 mg/L时对花粉萌发有轻微促进作用,当浓度增大时抑制作用较突出;2,4-D对花粉萌发存在明显抑制作用;当赤霉素、2,4-D、吲哚乙酸、萘乙酸、6-BA浓度分别达到50、30、30、15、25 mg/L时,花粉遭受浓度迫害出现畸形扭曲。吲哚乙酸、萘乙酸、赤霉素、2,4-D、6-BA对‘秦冠’花粉管生长和花粉萌发均无明显促进作用,并且随浓度增大抑制作用较明显,人工辅助授粉时用于添加剂效果不理想。     

纳米磁珠介导牡丹花粉转基因技术的初步分析

磁性纳米颗粒(magnetic nanoparticles,MNPs)可以吸附DNA,作为一种新型基因载体应用于转基因育种。本研究旨在探索MNPs介导外源基因进入牡丹花粉,为利用携有外源基因的花粉进行授粉,获得转基因种子提供依据,本研究制备了MNPs与DNA的复合物;利用扫描电子显微镜与透射电子显微镜观察了牡丹花粉的形态结构;用花粉离体萌发法测定了供试牡丹花粉磁转染前后的生活力。结果表明:MNPs与DNA的质量比为1:1~1:10时,MNPs均可以与DNA稳定结合,并可以保护DNA免受酶的消化作用;牡丹花粉为长球形,萌发孔为三孔沟类型,位于萌发沟内;‘凤丹’牡丹、‘京红’牡丹及紫斑牡丹的花粉均具有较高的萌发力,且经磁转染后的三种牡丹花粉萌发力保持在50%以上,能够用于正常杂交授粉。本研究首次采用四氧化三铁纳米磁转化技术介导牡丹花粉转基因,弥补了花粉介导牡丹转基因方法研究的空白,从而有助于牡丹分子育种进程的推进。     

转核酸酶barnase基因对小麦育性的影响

以小麦品系pm35为材料,利用基因枪法将核酸酶barnase基因导入小麦,获得12株阳性苗。至开花期,有7株表现为花粉明显减少,经花粉染色及花粉萌发率测定显示,该7株转基因小麦存在不同程度的花粉败育现象。进一步计算结实率发现,其结实率都明显下降,其中转基因植株10-11的结实率仅为3.43%。     

Slow-growing pollen-tube of colchicine-induced 2n pollen responsible for low triploid production rate in <Emphasis Type="Italic">Populus</Emphasis>

To reveal the mechanism of low triploid production, meiotic observation, induction of 2n pollen by colchicine, hybridization of colchicine-induced 2n pollen and pollen germination were conducted in Populus tomentosa clone 5088. Meiosis of the PMCs was initiated 24 h after it was cultured in the greenhouse. Results showed that the meiosis developed in a consecutive, asynchronous process. Analysis of the occurrence rate of colchicine-induced 2n pollen using the GLM-univariate revealed significant differences among the dominant meiotic stages (F = 45.822, P = 0.000). Significant differences also occurred among various colchicine injection times (F = 10.150, P = 0.004). The most effective stage for colchicine-induced 2n pollen occurred between the zygotene to diakinesis substages. Sixty-eight triploids were confirmed with both flow cytometric analysis and chromosome number-counting among the 3346 offspring. The highest triploid induction rate was 13.2%—significantly lower than the expected triploid production rate of 47.0%—indicating that the colchicine-induced 2n pollen was weak during fertilization. We found positive correlation between the efficiency of triploid production and the frequency of colchicine-induced 2n pollen (r = 0.961, P = 0.001) which suggests that the triploid production rate increased as the occurrence rate of colchicine-induced 2n pollen increased. Analysis using the GLM-univariate indicated that both pollen types (F = 87.50, P < 0.001) and durations of pollen germination (F = 7.61, P = 0.002) did have significant effects on the lengths of pollen-tubes. Haploid pollen-tubes were significantly longer than colchicine-induced 2n pollen-tubes. This suggests that haploid pollen has an advantage during fertilization.     

濒危植物珊瑚菜花粉生活力的测定

采用花粉离体萌发法,对珊瑚菜的花粉进行了生活力测定。结果表明:珊瑚菜初花期的花粉萌发率最高,为89.7%;在恒温25℃条件下,以0.6%琼脂+10%蔗糖+0.1%硼酸为离体培养基进行培养,有利于珊瑚菜花粉的萌发;低温（4℃）条件可延长珊瑚菜花粉的寿命,将其进行短期贮藏。     

The effects of high-temperature stress on the germination of pollen grains of upland cotton during square development

The reproductive stage of flowering plants is sensitive to high-temperature stresses. High temperature is a major factor influencing pollen grain viability in upland cotton (Gossypium hirsutum). The objective of this study was to identify the relationship between cotton pollen germination percentage and temperature by assaying the pollen germination of four upland cotton cultivars in vitro at different temperatures during the blooming period. The results showed that in vitro pollen germination percentage was related to the culture temperature of pollen germination and the temperature of the square development process. High temperature affected pollen development and germination, and high-temperature tolerance differed among the cotton cultivars. The pollen germination percentage decreased rapidly with changes in the culture temperature from 30 to 39 °C. A culture temperature of 35 °C might be a critical temperature for the pollen viability transition and could be used to screen cotton cultivars that have pollen grains with high-temperature resistance. Before the high-temperature stage, cultivars with rates of decrease in the percentage of pollen germination of less than 41 % at 35 °C relative to the rates at 30 °C might be considered as high-temperature tolerance cultivars, and cultivars with rates of decrease in the percentage of pollen germination greater than 41 % might be considered as susceptible cultivars. The high-temperature stress for pollen grain germination in vitro was greater than 30 °C, and the high-temperature stress for square development might be greater than 33 °C. Boll retention was significant; it was positively correlated with the pollen germination percentage and negatively correlated with temperature during the high-temperature stage. This study provided a method for rapidly screening cultivars (lines) with high-temperature tolerance pollen in upland cotton breeding.     

辣椒花粉生活力最佳测定方法的筛选

研究结果表明：I－KI染色法，醋酸洋红染色法，TTC染色法，过氧化物酶沉淀法和培养基离体萌发法等5种方法的适用测定现采花粉的生活力，而要测定贮藏花粉的生活力只能用TTC染色法和培养基离体萌发法。不论对何种来源的辣椒花粉进行生活力测定，以离体萌发法所得结果最为直观，稳定，是测定辣椒花粉生活力的最佳方法。     

白菜型油菜DFR基因的克隆与序列分析

By using RACE (rapid amplification ofcDNA ends) based homologous cloning strategy, we have successfully isolated the genomic and full-length cDNA sequences of a gene encoding typical DFR (dihydroflavonol-4-reductase) from black-seeded Brassica campestris L. var. oleifera DC.. The gene, designated BcDFR here, is 1 722bp in length and harbors 5 introns with typical splice sites of plant DFR genes. BcDFR cDNA is 1311bp in length with a 1 158bp ORF as well as a 25bp 5‘ UTR and a 128bp 3‘ UTR. The encoded BcDFR protein is 385 aa with a calculated Mw of 42.85kD and a pI value of 5.55. The nucleotide and amino acid sequences of this gene share extensive homologies to plant DFR genes of wide origins especially high similarities to Cruciferous DFR genes. Sequence analyses such as phylogenetic analysis, conserved domain search and substrate specificity region detection all indicated that BcDFR gene is a quite potentially biofunctional gene. Its cloning enables us to further dissect the possible relatedness between DFR gene and Brassica seed coat color traits and to create transgenic novel yellow-seeded rapeseed germplasm through antisense- or RNAi-suppression of DFR gene expression in black-seeded elite cultivars.     

电子束辐射对玉米及陆地棉花粉萌发率的影响

研究电子束辐照对花粉萌发率的影响,为电子束辐射诱变育种提供实验基础。对‘京糯28’玉米花粉和陆地棉花粉进行不同剂量的电子束辐照,在液体培养基中对辐照的花粉进行培养,观察并统计二者的花粉萌发率。玉米‘京糯28’花粉在200~600 Gy辐射剂量范围内花粉萌发率均高于对照组,其中以600 Gy辐射剂量的萌发率最高;而陆地棉在300~1000 Gy辐射剂量范围内花粉的萌发率均低于对照组,但在50 h后对其花粉进行培养时,各辐照组的花粉萌发率均高于对照组。电子束辐照剂量在200~600 Gy范围内对‘京糯28’玉米花粉萌发率有促进作用,在300~1000 Gy范围内对陆地棉花粉萌发率有不同程度的抑制作用,但在一定程度上延长了陆地棉花粉生活力。     

Combining ability of low phytic acid (<Emphasis Type="Italic">lpa1</Emphasis>-<Emphasis Type="Italic">1</Emphasis>) and quality protein maize (QPM) lines for seed germination and vigour under stress and non-stress conditions

Grain yield is more likely to be compromised by poor seed germination and vigour in low phytic acid (LPA) and quality protein maize (QPM) than normal maize (Nm), especially when grown under stressful tropical environmental conditions. The objectives of this study were to determine the effect of stress conditions on seed germination and vigour traits (percentage germination, seedling dry weight, average root and shoot length and vigour index) and to determine the GCA effects of the parental lines and SCA effects of the crosses. A ten parent half diallel (LPA, Nm, QPM lines) was subjected to the standard germination (non-stress) and accelerated aging (AA) (stress) tests. All seed lots were at the same physiological age and produced under the same season and conditions. Genotypic and group differences were investigated. General combining ability (GCA) and specific combining ability (SCA) effects were significant (P ≤ 0.001) for all traits under both stress and non-stress conditions indicating that both additive and non-additive gene effects were significant. Generally SCA effects were superior for all traits. The LPA lines displayed 61 % reduction in germination and 23–52 % reduction in vigour under stress conditions thereby underscoring challenges that are expected in deploying LPA maize in stress conditions. However, LPA line CM 31 exhibited large positive and mostly significant GCA effects, while two LPA × LPA crosses showed significant positive SCA effects. Results indicate breeding is required to improve both germination and vigour of the LPA lines to adapt them to tropical conditions that are generally stress-prone.     

一个棉花β-1,3-葡聚糖酶基因全长cDNA的克隆与特征分析

利用本实验室分离的防卫基因类似物片段,通过5’和3’RACE,从海岛棉海7124中克隆了一个β-1,3-葡聚糖酶基因全长cDNA,命名为GbGLU。其全长1 266 bp,含一个编码361个氨基酸残基的开放阅读框。该基因编码产物理论分子量和等电点为MW = 39.66 kD,pI = 9.15,含有糖基水解酶家族17的保守结构域,N端有26个氨基酸残基组成的信号肽。Southern结果显示,GbGLU在棉花基因组中以单拷贝形式存在,研究发现,GbGLU的表达受黄萎病菌的诱导。聚类分析表明,棉花葡聚糖酶基因可分为3类,分别含有糖基水解酶家族3、9、17的保守结构域。无论棉花中的还是本研究中参考的其他作物中,受病菌诱导的葡聚糖酶基因都含有糖基水解酶家族17的保守结构域,故推断其为此类基因参与抗病反应的功能域。