菠萝微卫星指纹图谱的构建

初步构建了菠萝种质的SSR指纹图谱,为菠萝种质鉴定提供了依据.从菠萝EST-SSR开发的20对微卫星引物扩增31份不同的菠萝种质,筛选出能够稳定扩增且带型差异明显的SSR引物.挑选5对多态性高、分辨率高、重复性好的SSR引物对,作为标准引物对材料进行扩增.每对引物得到的多态性带数8~15个,平均为11条;引物的多态信息...     

Factors affecting mango (Mangifera indica L.) protoplast isolation and culture

The major factors influencing protoplast isolation and culture of mango (Mangifera indica L.) cv. Kensington Pride were investigated. The resultant protocol was used to compare plating efficiency among 4 mango cultivars. Most responses differed between proembryonic masses (PEMs) and leaf sources. Protoplast yields of 15.22 × 10⁶ g⁻¹ from PEMs and 8.68 × 10⁶ g⁻¹ from greenhouse-derived leaves were obtained in a solution of 0.7 M mannitol CPW plus 1.5% cellulase, 1% hemicellulase and 0.75% macerozyme for PEMs or 0.5 M mannitol CPW plus 1.5% cellulose, 1% hemicellulase and 1.5% macerozyme for leaves. Culture in Ca-alginate beads with initial plating densities (IPD) of 2.5 × 10⁴ Pp mL⁻¹ for PEMs and 2.5 × 10⁵ for leaves gave the highest plating efficiencies (FPE). For PEMs 1 mg L⁻¹ 2,4-d and 3.5 mg L⁻¹ kinetin gave an FPE of 2.85% whereas lower kinetin (2 mg L⁻¹) plus 0.5 mg L⁻¹ 6-BAP was most effective for leaves (FPE of 2.12%). Most protoplast mortality occurred during the first week of culture and was more severe in liquid culture. In Ca-alginate beads, protoplast survival at 14 days was higher for PEMs (30%) than leaf (21%) as was the frequency of cell division (17.6% compared to 13.6%). PEMs protoplasts continued development through embryogenesis to in vitro plantlet regeneration whereas leaf protoplasts underwent cell division up to 40-cell colonies but failed to proceed further. For PEMs, polyembryonic cvs. Kensington Pride and Keow Savoey produced higher FPE (1.95%) than monoembryonic cvs. Tommy Atkins and Keitt (1.75%). There was no effect of cultivar for leaf protoplasts.     

从菠萝薄细胞层切片直接诱导体细胞胚

 以菠萝(Ananas comosus L. Merrill) 杂交种MD1 无菌苗为材料, 通过培养薄细胞层切片 (Thin cell layer, TCL) 的方法, 研究直接再生胚的固体培养体系。结果证明, 预培养的设计2 (在预培养基PT上黑暗预培养无菌苗1周后, 从预培养苗茎顶端切取TCL, 在MS + 2, 4-D 2 mg·L ^{- 1}或1 mg·L ^{- 1}胚诱导培养基EI2上黑暗培养1周, 然后转到光照条件下继续诱导胚1周, 再在成熟胚培养基EM上培养2周, 最后转入幼苗培养基PL上培养) 是直接诱导获得体胚的最佳途径; 诱导的体胚最初均来自于薄细胞层上微管束附近的细胞; picloram有利于芽的诱导, 2, 4-D有利于直接再生胚, 但如果球形胚形成后一直培养在较高浓度的2, 4-D培养基中, 将阻碍球形胚和心形胚进一步发育为成熟胚, 反而促进芽的生成。     

Heterosis studies in chilli (Capsicum annuum L.)

By top-crossing 12 different homozygous chilli varieties with a genetic male sterile (MS), 12 hybrids were produced. Heterosis over mid-parent (MP) and superior parent (SP) was studied for days to flower, plant height, number of primary branches, length of fruit, number of fruits per plant and fruit yield per plant. Degree and direction of heterosis varied greatly for different characteristics and crosses studied. The hybrid MS × K-2 exhibited significant positive heterosis for fruit yield over its SP (157%), followed by MS × Pant-C-1, MS × Japanese bunch variety and MS × Bhiwapur local, indicating that male sterility can be utilised for commercial exploitation of hybrid vigour. Heterosis was observed for days to flower, plant height, number of primary branches, fruit length and number of fruits per plant.     

Identification of reference genes for real-time quantitative polymerase chain reaction based gene expression studies on various Olive (Olea europaea L.) tissues

Reference genes are essential for the normalisation of the expression data of quantitative real-time PCR for the purposes of validation. Although several reference genes have been validated for the olive, comprehensive analyses including an excessive number of candidate reference genes still require study in various olive tissue samples. In this work, a total of 40 candidate reference genes were tested for their stability in 8 different olive tissues (root, apical bud, lateral bud, pedicel, young leaf, mature leaf, fruit mesocarp, and seed) with the utilisation of the most popular software programs including GeNorm, NormFinder, BestKeeper, and ΔC_t. The analyses of expression stability of candidate reference genes using quantitative real-time PCR demonstrated ubiquitin-conjugating enzyme (UBC1) as the most stable reference gene for the studied tissues of the olive. The GeNorm software also calculated the optimum reference gene combinations as two which consist of UBC1 and the Clathrin adaptor complex medium subunit (CLATHRIN) genes. This study provides the most stable reference gene combination for normalisation of target genes for quantitative real-time PCR gene expression studies on the olive.     

香蕉原生质体培养和体细胞杂交研究进展

香蕉是继水稻、小麦、玉米之后的世界第4大粮食作物和近5亿发展中国家人口的主要粮食,香蕉产业的健康发展具有重要的经济和社会地位。但由于香蕉品种的高度不育性和多倍性,制约了用传统育种方法培育生产实践中所需抗病虫害新品种。通过原生质体融合进行的体细胞杂交育种有望克服这些障碍。原生质体培养技术是细胞融合所必需的基础技术之一,自1984年首次进行香蕉原生质体分离研究以来,有关香蕉原生质体分离、培养和融合等方面的研究陆续开展,相继有超过10个品种成功获得再生植株,并且开始了体细胞杂交研究的探索。作者就香蕉原生质体培养和体细胞杂交研究进展进行综述。     

Cloning,characterisation, and expression analysis of an oleosin gene in coconut (Cocos nucifera L.) pulp

Summary

Oleosins are structural proteins found in oil bodies, organelles found in the cells of plant tissues with a high oil content that undergo extreme desiccation as part of their maturation process. Oleosins stabilise oil bodies. In this paper, a full-length cDNA sequence homologous to oleosin, a seed-storage oil-body protein, from coconut (Cocos nucifera L.) was identified from cDNA libraries during fruit development and characterised. The gene, termed Coco-Ole, contained an open reading frame of 375 bp encoding a polypeptide of 125 amino acids. The predicted amino acid sequence of coconut oleosin had a molecular mass of 13.0 kDa, and showed 92% (AAF76238.1) and 67% (AAC02239.1) sequence similarity to oil palm (Elaeis guineensis Jacq.) and rice (Oryza sativa) oleosin proteins, respectively. The amino acid sequence clustered in the same branch as oil palm in the cladogram, but was distant from other species. The results of semi-quantitative RT-PCR indicated that the Coco-Ole gene was expressed only in the pulp, and its expression increased significantly during pulp development. Compared with fluctuations in oil content, expression of the Coco-Ole gene was consistent with the anabolism of oil during pulp development. The cloning and sequencing of the Coco-ole gene provides a new marker for studies on oil body biogenesis and fruit development in coconut.     

A recirculating hydroponic system for studying peanut (Arachis hypogaea L.)

Peanut (Arachis hypogaea L.) plants were grown hydroponically, using continuously recirculating nutrient solution. Two culture tray designs were tested; one tray design used only nutrient solution, while the other used a sphagnum-filled pod development compartment just beneath the cover and above the nutrient solution. Both trays were fitted with slotted covers to allow developing gynophores to reach the root zone. Peanut seed yields averaged 350 gm-2 dry mass, regardless of tray design, suggesting that substrate is not required for hydroponic peanut production.     

Poinsettia protoplasts - a simple, robust and efficient system for transient gene expression studies

ABSTRACT: BACKGROUND: Transient gene expression systems are indispensable tools in molecular biology. Yet, their routine application is limited to only few plant species and often requires substantial equipment and facilities. The high content of chloroplasts and chlorophyll can impede downstream applications of transformed cells from green plant tissue. RESULTS: We describe a fast and simple technique for the high-yield isolation and efficient transformation (>70%) of mesophyll-derived protoplasts from red leaves of the perennial plant Poinsettia (Euphorbia pulccherrima) for which no particular growth facilities or expensive equipment are needed. Poinsettia protoplasts display an astonishing robustness and can be employed in all commonly-used downstream applications, such as subcellular localisation (multi-colour fluorescence) or promoter activity studies. Due to a low abundance of chloroplasts or chromoplasts, problems encountered in other mesophyll-derived protoplast systems (particularly autofluorescence) are alleviated. Transgene expression is detectable within 90 min post-transformation and lasts over several days. CONCLUSIONS: The simplicity of isolation and transformation renders Poinsettia protoplasts an attractive system for transient gene expression experiments, including multicolour fluorescence, subcellular localisation and promoter activity studies. In addition, they offer hitherto unknown possibilities for anthocyan research and industrial applications.     

The modification of sex expression in papaya (Carica papaya L.)

Field trials with six times normal nitrogen supply produced a significant increase in the proportion of female papaya plants. The addition of nitrogen to the soil increased the leaf nitrogen content but had no effect on the amino acid content. Leaves of male plants contained more nitrogen than female plants but no difference could be detected in the contents of amino acids in the leaves and flower buds. Foliar application of 1-naphthalene acetic acid (NAA) and 2-chloroethyl-trimethyl ammonium chloride (CCC) resulted in a higher female : male ratio, but maleic hydrazide (MH) and gibberellic acid (GA₃) enhanced maleness. The animal sex hormone stilboesterol dipropionate had no apparent effect on sex expression but the application of testosterone propionate increased the proportion of male plants.     

Further studies on graft-induced off-season flowering and fruiting in mango (Mangifera indica L.)

Summary

Defoliated shoots of cvs Alphonso, Dashehari, Totapari etc. (receptors), could be induced to flower within four weeks during the off-season by veneer grafting them to leafy shoots of the off-season flowering cv. Royal Special (donor) during the non-flowering season. Experiments on defoliating the donor and receptor shoots revealed the crucial dual role of the leaves in flowering. In the ‘floral cycle’, leaves from the donor promoted flowering, whereas leaves on the receptors, in the vegative phase, were inhibitory, and prevented graft-induction of the receptors. Thus, for graft induction, leafy donors and defoliation of receptors were essential. The inhibitory effect of the leaves on the receptors was localized and did not affect flowering of the donor shoots’ The similarity between these findings and those in the herbaceous, day-length sensitive species strengthens the view that flower formation is controlled in the same way in herbaceous and in woody perennial species. A minimum threshold of the floral stimulus appeared to be another requirement for an optimum flowering response. This was concluded from an experiment in which the leafy receptors were defoliated on different dates during the off-season flowering cycle. At the end of cycle, the defoliated receptors produced small panicles, and finally only vegetative shoots, probably indicating sub-threshold levels of the stimulus. Bud activity at the apex in the receptor was also an important pre-requisite for graft-induction. Some veneer-grafted scions which remained dormant during the flowering cycle of the donor and which sprouted much later after the completion of off-season flowering, ‘escaped’ the stimulus and invariably turned out to be vegetative. It is postulated that the cyclic synthesis of the floral stimulus in the leaves in an inductive cycle, and the gap between two such cycles, mainly decides the flowering behaviour of mango cultivars—biennial, annual and multiflowering. Two other requiremets are the absence of non-induced leaves and the synchronization of meristematic activity in the bud with the inductive cycle. Juvenile shoots from one to four year old seedlings could not be graft-induced—unlike the shoots from mature, six year old seeding trees. This juvenility effect continued into the second year when defoliation of the receptor shoots had no effect and failed to induce them to flower.     

拟南芥叶绿体SSR引物在甘蓝上的应用

 根据拟南芥的叶绿体全基因组序列,搜索获得89个叶绿体基因组SSRs（cpSSRs）,平均每1.736 kb出现1个SSR。针对这些SSR设计cpSSR引物58对,其中32对能在甘蓝上应用。5对引物在甘蓝C300、C322和波里马油菜N478、N479间有多态性,这些片段均位于基因内含子区或基因间隔区。测序结果显示单核苷酸A/T重复数从8个到13个不等。同源性比较表明C300、N479之间的同源性大于两者与拟南芥之间的同源性。     

普通杏品种SSR遗传多样性分析

利用来自杏、桃和扁桃的25对SSR多态性引物对66个普通杏品种进行了PCR扩增及变性聚丙烯酰胺凝胶电泳,共检测出284个等位位点,每对引物的等位位点数在3～17之间,平均为11.36。通过NTSYS软件计算得到的Dice相似性系数为0.083～0.987,平均值为0.370,表明中国普通杏种质资源的遗传多样性丰富。UPGMA法聚类将66份材料分为5个组。SSR标记反映出的品种间亲缘关系与根据地理生态类型对杏种质的分类结果基本一致。来自四川和贵州的多数品种独立聚成一组,表现出与其它品种较远的亲缘关系,该地区可能存在特异性较高的种质资源。证实了扁桃基因组SSR引物可用在杏SSR标记的研究中。     

Effect of short-term exposure to high-temperature on total gene expression in the leaves of four raspberry (Rubus idaeus L.) cultivars

Summary

The effect of high temperature stress (27ºC or 37ºC for 24 h) on total gene expression profiles in the annual-fruiting raspberry (Rubus idaeus L.) cultivars ‘Autumn Bliss’, ‘Autumn Treasure’, ‘Erika’, and ‘Polka’ were evaluated at the floral initiation stage using a customised Rubus microarray. Significantly affected genes were obtained by pairwise t-tests using ‘volcano plots’ for each cultivar × treatment. A 10ºC elevation in temperature altered levels of expression, in at least one cultivar, of 644 differentially expressed genes in total, with ‘Erika’ and ‘Autumn Treasure’ showing elevated expression of 38 genes compared to ‘Autumn Bliss’ and ‘Polka’. We identified 12 common candidate genes that were modulated differentially in ‘Autumn Bliss’ and ‘Erika’ at 37ºC compared to 27ºC. In addition, two aquaporin genes (PIP1 and TIP2) were down-regulated in ‘Autumn Bliss’, but up-regulated in ‘Autumn Treasure’, ‘Polka’, and ‘Erika’ at 37ºC. Other down-regulated genes from the list of 38 genes included those encoding major latex-like proteins, plasma membrane proteins, cysteine rich proteins, and other stress-related proteins. Validation by real-time quantitative RT-PCR (RT-qPCR) indicated subtle changes in differential gene expression, suggesting a mild response to heat stress. This study used molecular tools to increase our understanding of, and to identify candidate genes involved in, the heat stress response of four annual-fruiting raspberry cultivars.     

西瓜果实长圆的遗传及候选基因定位

<正>目的与意义:果实形状作为一个非常重要的农艺性状,在许多作物中已经有了非常深入的研究,同时也获得了许多控制果实形状的候选基因。随着2012年西瓜全基因组测序的完成,西瓜分子生物学的研究得到了快速发展,目前关于西瓜遗传图谱的报道有很多,根据遗传图谱也得到了许多性状的QTLs,尤其是西瓜果实的长度、宽度、果形指数等。然而目前还没有关于西瓜果实形状候选基因定位的报道。因此笔者通过配制杂交组合,解析西瓜果实形状