Low temperature storage and in vitro germination of cherimoya (Annona cherimola Mill.) pollen

Due to the protogynous dichogamy of cherimoya and to the absence of proper pollinating vectors, hand-pollination with fresh pollen is a common practice for cherimoya commercial production. In order to optimize the process of hand-pollination, in this work we have studied the conservation of cherimoya pollen at −20, −80 and −196 °C for up to 3 months. In vitro pollen germination of fresh pollen was 57.1% and it was progressively reduced with conservation time at the three temperatures studied reaching a minimum after 3 months of storage of 10.4%, 14.2% and 13.6% at −20, −80 and −196 °C, respectively. Differences in germination among temperatures were only significant during the first 2 weeks of storage. Field pollinations with pollen stored for up to 3 months at the three temperatures show no yield differences compared to pollinations performed with fresh pollen. The results indicate that pollen collected and stored at sub-zero temperatures at the beginning of the cherimoya blooming season can be used along the whole blooming season avoiding the need of collecting fresh pollen daily.     

梨组织中苹果褪绿叶斑病毒的原位RT-PCR检测

 为了建立梨树苹果褪绿叶斑病毒原位RT-PCR检测技术, 用已知带有苹果褪绿叶斑病毒的库尔勒香梨和无病毒实生苗叶片为材料, 利用D IG标记, 研究了香梨病毒的叶片石蜡切片IS-RT-PCR检测技术。包括AMV逆转录酶、dNTPs、RNasin及互补引物浓度对cDNA合成的影响, 退火温度、Taq DNA聚合酶、Mg²⁺ 、引物浓度及循环次数对原位PCR效果的影响。结果表明: RNasin的用量大于0.2 U·μL^-1时,信号强度随着RNasin量的加大而增强; 只有当dNTPs浓度达1.0 mmol·L^-1时才能生成一定量的cDNA;AMV浓度在0.3～0.5 U·μL^-1均可进行正常的逆转录, 而且在该范围内产物的量随AMV浓度提高而增多; 引物浓度达到0.9μmol·L^-1以上时才能进行有效的逆转录, 并且生成的cDNA的量随引物浓度增大而增加。原位扩增ACLSV的cDNA适合退火温度为56℃。循环20～30次可出现较强的蓝色信号, 引物浓度在0.8～1.2μmol·L^-1时显色较好; Taq酶浓度为20 U·mL^-1以上, 均显示较深的蓝色; Mg^{2 +}浓度为1.5mmol·L^-1就可满足原位PCR所需。获得了苹果褪绿叶斑病毒的原位PCR优化检测体系, 利用建立的优化程序对香梨样品进行了检测验证, 取得了很好效果。     

不同培养方法对昆虫病原线虫质量影响及其评价

采用传统的White trap法、Lowtek培养法、液体和固体培养4种方法繁殖异小杆线虫[Heterorhabditis bac-teriophora(Hb-1)]和斯氏线虫[Steinernemacarpocapsae(Sc-2)],测定和计算了线虫的产量、质量和成本。试验结果显示,4种方法培养对线虫侵染力和存活率有一定影响,但差异不显著(p<0.05),2种线虫从高到低的顺序为:White trap法>Lowtek法>固体培养>液体培养;产量最高的是固体培养法,其次是液体培养法,Lowtek法最低;繁殖成本从低到高的顺序为:Lowtek法<固体培养     

海南坡鹿迁地保护现状与建议

迁地保护的海南坡鹿种群可以分为野放、半野放和圈养等3种主要类型。野放种群个体数量最多，约占迁地保护种群个体总数的55．3％；其次为半野放种群，其个体数量占35．6％；圈养种群个体数量最少，只占9．1％。半野放是一种较为稳妥的坡鹿种群恢复方式。在海南坡鹿的迁地保护中，应该增加半野放种群个体数量。本文就海南坡鹿的迁地保护提出了7点保护管理及科学研究建议。     

Genetic analysis of an attenuated <Emphasis Type="Italic">Papaya ringspot virus</Emphasis> strain applied for cross-protection

Papaya ringspot virus (PRSV) HA 5-1, a nitrous acid-induced mild mutant of severe strain HA, widely applied for control of PRSV by cross-protection, was used to study the genetic basis of attenuation. Using infectious clones, a series of recombinants was generated between HA 5-1 and HA and their infectivity was analyzed on the systemic host papaya and the local lesion host Chenopodium quinoa. The recombinants that contained mutations in P1 and HC-Pro genes caused attenuated infection on papaya without conspicuous symptoms, similar to HA 5-1. The recombination and sequence analyses strongly implicated two amino acid changes in the C-terminal region of P1 and two in HC-Pro of HA 5-1 involved in the attenuated infection on papaya. The recombinants that infected C. quinoa plants without local lesions contained the same mutations in the C-terminal region of HC-Pro for attenuated infection on papaya. We conclude that both P1 and HC-Pro bear important pathogenicity determinants for the infection on the systemic host papaya and that the mutations in HC-Pro affecting pathogenicity on papaya are also responsible for the inability to induce hypersensitive reaction on C. quinoa.     

花椰菜未受精子房、花托、花序轴离体培养诱导成苗研究

以花椰菜的花托、花序轴和未受了房为外植体，诱导出不定芽，并进行继代增殖培养，在工增殖培养基上每隔２５～３０天，不定芽可以增殖５～６倍，不加激素的１／２ＭＳ培养基上，试管苗生根率达到９８％以上，移栽成活率达到９５％以上。经观察，花托和未受精了再生植株是由外植体直接分化而来，不经过愈伤组织阶段，这样可避免不良变异，保持材料的遗传稳定性。     

苹果叶片离体培养中秋水仙素加倍效应的研究

用不同浓度的秋水仙素溶液处理苹果的栽培品种嘎拉试管苗的离体叶片，发现以０．５％的秋水仙素溶液处理４ｄ效果最佳，诱变频率达５６．１％。诱变后获得的四倍体植株在形态学和细胞学上均发生了明显变异。与对照株比较，变异株茎矮、粗壮、节间短、叶片厚、叶色浓绿；叶背面的气孔体积增大，单位面积气孔数目减少、茸毛数量增加；染色体数目加倍，２ｎ＝４ｘ＝６８（对照２ｎ＝２ｘ＝３４），细胞核核仁数目增加。梢端组织学切片证明，变异株中同质突变体的频率达７３％。     

Development of immunocompetence of broiler chickens

Subpopulations of T-cells, B-cells, macrophages and ellipsoid-associated reticular cells (EARC) could be demonstrated by immunohistochemical staining early in the development of chicken spleen. However, the typical structures of the spleen, such as the peri-arteriolar lymphoid sheath (PALS) and the ellipsoids with their surrounding ring of macrophages, were only formed around embryonic day (ED) 20. These structures and especially the B-cell compartment, i.e., the peri-ellipsoid lymphoid sheath (PELS) gradually matured during the first week posthatch.

Therefore, we analysed at what age broiler chickens could generate a humoral response against the thymus-dependent antigen bovine serum albumin (BSA). Chickens were immunised in ovo (ED16 and ED18) and at 1, 7 and 12 days of age and subsequent BSA-specific immunoglobulin (Ig) M and IgG responses were measured up to 10 days postimmunisation (DPI). No major differences were observed in the relative growth rates, while hatchability was only slightly reduced. Only in chicks immunised on 12 days of age, IgM and IgG responses were high with a normal kinetic pattern. In chicks immunised on 7 days of age, responses were just detectable, but they were absent in chicks immunised in ovo and on the day of hatching (Day 1).

In a subsequent experiment, 1-, 7- and 12-day-old chicks were BSA-immunised and Ig responses were measured for a longer period up to the age of 28 days. The IgG response of chicks immunised at 1 day of age was lower and occurred later (from 28 DPI) than the response of chicks immunised at 7 and 14 days of age (from 14 DPI). It was not increased by a booster immunisation on 29 days of age, in contrast to the response of chicks immunised at 7 and 14 days of age. These findings indicate that vaccination at 1 day of age does not activate the B-cell response resulting in antibody production and support the idea that the immune function of the late embryonic and neonatal chickens is not entirely developed due to the incomplete structural organisation of their secondary immune organs.     

Micropropagation of Dendrobium draconis Rchb. f. from thin cross-section culture

An efficient procedure is outlined for rapid and mass in vitro propagation of an orchid, Dendrobium draconis Rchb. f. through in vitro culture of thin cross-sections (TCSs) derived from young stems. The TCS explants were excised along the stem from the base to shoot tip of 6-month-old plantlets and cultured on Murashige and Skoog (MS) medium supplemented with 20 g/l sucrose and different concentrations of N⁶-benzyladenine (BA), kinetin (Kn) and 1-naphthaleneacetic acid (NAA), either individually or in combination. Protocorm-like bodies (PLBs) were directly induced from the TCS explants and completely developed into shoots within 6–7 weeks. The optimal growth regulators combination for maximal PLB development was 2 mg/l BA and 1.0 mg/l NAA, giving rise to 68% of responding explants with an average 11 PLBs per explant. Shoot development was best achieved on MS medium containing sucrose and coconut water. Plantlets, 6–8 cm height were transplanted into coconut husk peat with 92% survival rate in a nursery.