无线遥测和刺激技术建立兔房颤模型的探索

目的探索一种在无线遥测和刺激技术基础上的兔房颤模型的制作。方法新西兰兔皮下植入自主研发的植入式遥测刺激器,植入式遥测刺激器的制作是以TI公司（德州仪器）的MSP单片机和TI公司的RF无线收发芯片CC2250为核心开发设计。优化植入系统设计以满足新西兰兔房颤模型建立的探索实验;植入子植入新西兰兔腹部皮下,采集电极留置于左上肢和右上肢腋下皮下,两个刺激电极分别缝合于左心耳和左心房上,通过无线收发采集和刺激信号;实现利用Powerlab生理记录仪连续监测体表I导联心电信号,并通过专用计算机程序刺激软件,发放间歇（刺激2 s,暂停2 s）高频（频率20 Hz）阈上（强度2 mA,脉宽1 ms）刺激,若间歇期内出现房颤,则人为干预中止刺激,若转为窦性心律,则继续刺激。结果植入式遥测刺激器在体内可稳定工作（包括采集模拟心电信号和发放刺激）30 d,植入新西兰兔体内刺激3周后可诱导出房颤,持续时间〉48 h。结论用新西兰兔代替比格犬建立基于无线遥测和刺激基础上的房颤模型是完全可行的,同时也体现了动物福利优化和替代原则。     

运动后心力及心率同时遥测的心力恢复趋势与心率恢复趋势的对比研究

为了更加全面地研究和评估心肌的变时性和变力性,研制了能够在运动场地同时采集心力和心率的心音信号遥测系统。采集了50名体育系学生和30名其他系学生在完成规定运动量的登梯运动后,5min内连续变化的心音信号。对采集的数据经过3次样条插值、均匀采样和小波变换滤波后,得到了平滑的心力恢复趋势曲线和心率恢复趋势曲线。统计分析表明,大负荷运动量下的心脏储备主要来自于心力储备而不是心率储备(P<0.001),和其他系学生相比,体育系学生具有的高水平心功能不仅在于具有更大的心力储备指数(P<0.001),而且还在于具有更快的心力恢复速度(P<0.001)。同时检测心力恢复趋势与心率恢复趋势会有益于全面评估心脏功能。     

同步遥测分析大鼠棕色脂肪产热与体温昼夜节律变化的时间关系

目的：同步遥测棕色脂肪组织（BAT）产热与体核温度昼夜律变化的时间曲线,分析二者昼夜节律变化的时间关系。方法：实验用成年雄性SD大鼠,在22℃环境温度下,明暗时间各12h,昼光时间为06：00h-18：00h,同步无线遥测体核温度（TC）、BAT温度（T队T）、腋窝温度（Tax）和动物活动的昼夜节律变化。结果：①在昼光中,TBAT较TC低0．67％,而在暗光中二者则相似。大鼠从昼光进入暗光时,TBAT升高的速率较TC升高速率快,开始上升的时间较TC提前8min;而从暗光进入昼光时,TBAT开始下降的时间则较TC提前4min。②Tax的昼夜节律幅度与TC相似,但无论动物在明光期或暗光期中,Tax均低于同步测量的TC。③从昼光期转入暗光期时,动物的行为活动出现增加反应先于TBAT和TC开始上升的时间。结论：实验结果证明,在暗光期中大鼠TC升高与BAT产热增加有关,说明BAT昼夜节律性产热的变化在维持体温昼夜生理节律中有重要的作用。     

丙型肝炎病毒多表位抗原基因的构建与免疫原性研究

丙型肝炎病毒(HCV)基因易发生变异, 尤其是含中和抗原表位的高变区1(HVR1)变异性最大. 模拟HVR1的B细胞表位具有涵盖多种天然表位的抗原特性, 保守的T细胞表位具有各型间的相对保守性. 为解决HCV高变性造成的疫苗研究障碍, 我们选取HCV E2区HVR1(384～410 aa)模拟B细胞表位9条、C区的保守CTL表位2条(35～44 aa, 132～140 aa)、NS3区保守的CTL表位1条(1073～1081 aa)及NS3区保守的Th表位1条(aa 1251～1259), 各表位之间以插入3个氨基酸为连接臂, 人工合成上述13条表位基因串联的HCV多表位抗原基因(mfc). 将mfc与gst基因融合, 表达了多表位抗原蛋白GST-MFC. 同时, 构建了白介素-2信号肽基因、PADRE表位基因和mfc基因串联的候选HCV DNA疫苗, 即质粒pVAX1.0-st-mfc. 以GST-MFC蛋白免疫家兔和质粒pVAX1.0-st-mfc免疫小鼠, 采用ELISA和Western blot方法, 应用10条具有代表性的HCV HVR1合成肽进行检测, 证明有9条HVR1合成肽能与所有免疫动物血清反应, 交叉反应率(cross reactivity, CR)为90%. 应用HCV抗体阳性的感染者血清与多表位抗原GST-MFC蛋白进行反应, 证明其反应识别率(reactivity frequency, RF)为75%. 上述结果表明, 筛选合成的HCV多表位抗原基因mfc具有HCV中和抗原表位的特征, 可作为HCV疫苗研制的候选基因.     

药物传输系统(DDS)的研究现状与发展前景

药物传输系统(drug delivery system,简称DDS)的出现大大改善了药物的药理学和药效学特性,目前已有一些抗肿瘤和抗真菌的DDS制剂应用于临床。基因组学和蛋白质组学的发展为制备新药提供了契机,如能最大限度地利用DDS系统的优势并将其应用于新药的体内传输及受体靶向治疗方面,将推动新药以至医学的进一步发展。     

高山峡谷地区无线电遥测与GPS 空间定位的比较:野外放归大熊猫的跟踪定位

以佩戴具有无线电发射功能的GPS 颈圈(Lotek GPS_ 4400M) 的放归大熊猫“祥祥”作为目标动物, 2006 年4 月至2007 年2 月,采用无线电遥测技术(RT)和GPS 跟踪技术在卧龙自然保护区的“五一棚”区域, 每日监测大熊猫在野外环境下的生存状况、移动规律和觅食行为。为了比较RT 和GPS 在高山峡谷地区空间定位的可行性和有效性,我们引入空间定位率、地形特征、空间定位差、巢域大小和日移动距离等指标来分析RT 和GPS 之间的定位差异。结果表明:RT 的空间定位效率明显高于GPS 的自动定位(P < 0. 001),分别是54.1%(绘图法)和45. 2% (≧ 2D);不同月份RT 和GPS 的空间定位率之间具有显著性差异(P < 0. 05),这与大熊猫不同月份的海拔活动范围和觅食行为特性密切相关。RT 位点的地形指数中坡度高于GPS,坡向和海拔高度较GPS 定位点低,两种无线电遥测方法(两点直接计算法和绘图法)之间没有显著性差异(P > 0.05);同一天位点之间的距离(空间定位差)平均450 ～ 660 m 左右;RT 与GPS 所估测的大熊猫巢域大小,除5 月、9 月和12月RT 低于GPS 外,其余月份为前者高于后者,但无显著性差异(P > 0. 05);日移动距离除12 月份RT 小于GPS 外,其余月份都呈现出RT 大于GPS 的格局,统计检验结果两者之间差异显著(P < 0. 05);两种无线电遥测方法所测指数之间都无明显差异(P > 0. 05)。这说明RT 遥测和GPS 定位都可以应用于高山峡谷地区野生动物的生态学研究,而且GPS 无线电颈圈在亚高山和高山森林中具有可行性和有效性。     

抗禽流感病毒多表位DNA疫苗的构建及其免疫效力研究

多表位DNA疫苗是建立在常规DNA疫苗基础上的一种新型疫苗。它是用表位作免疫原,这样就比较容易在一个表达载体上克隆病原体的多个抗原基因中具有免疫活性的部分。本试验以H5N1亚型禽流感病毒的HA和NP基因及其表位为基础构建了4个重组质粒：1 pIRES/HA（表达全长的HA基因）;2 pIRES/tHA（只表达HA基因的主要抗原表位区）;3 pIRES/tHANpep（融合表达HA基因的抗原表位区和NP基因的3个CTL表位）;4 pIRES/tHANpep-IFN-γ（用鸡的IFN-γ基因取代质粒pIRES/tHANpep中的neo基因）。分别用这4个重组质粒和空载体质粒pIRES1neo肌注免疫30日龄SPF鸡。免疫3次,间隔为2周,每次每只鸡的剂量为200μg。第3次免疫后两周以高致病性禽流感病毒H5N1强毒攻击,免疫及攻毒前后均采血检测HI抗体效价和外周血CD4⁺、CD8⁺T细胞的变化。结果发现,攻毒前各质粒免疫组均检测不到HI抗体,攻毒后1周存活鸡HI抗体效价迅速升高到64～256。流式细胞仪检测显示外周血CD4⁺、CD8⁺T细胞在疫苗免疫后都有不同程度的升高。空载体质粒对照组鸡（10只）在攻毒后3～8 d内全部死亡,其他各重组质粒免疫组鸡都获得了部分保护,保护率分别是：pIRES/HA组为545%（6/11）,pIRES/tHA组为30%（3/10）,pIRES/tHANPep组为36.3%（4/11）, pIRES/tHANPepIFNγ组为50%（5/10）。这些结果表明我们构建的多表位DNA疫苗能够诱导机体产生特异性免疫应答,并在同型禽流感强毒攻击时对鸡只提供了一定的保护。     

动物snmRNA的系统识别与鉴定

小分子非编码RNA(snmRNA)在调控基因的转录和转录后加工、细胞分化和个体发育、遗传和表观遗传等几乎所有的重要生命活动中发挥关键作用。建立和发展snmRNA研究技术,系统地发现和注释基因组中的snmRNA基因并阐明其生物学意义是当前RNA组学的首要任务。围绕snmRNA的系统识别与鉴定等问题,该文对近年来采用实验技术和计算机预测方法发掘snmRNA所取得的主要研究成果进行综述。     

神经元信息编码的代谢消耗：动作电位与能量效率

人脑是一个高效、可靠的信息处理系统,它主导着个体的认知、情感、意识与行为,这些功能的实现需要不断地消耗代谢能量.大脑的能量需求主要被神经元信息编码所消耗,相应的亚细胞过程包括产生和传导动作电位、维持静息电位以及突触传递.神经元编码信息的主要载体是动作电位序列,它的产生与传导贡献了大脑的大部分代谢消耗.动作电位的能量消耗受离子通道的生物物理特性控制.生物物理特性的细胞特异性和空间异质性使得动作电位对代谢能量的利用效率呈现高度可变性,它为理解神经元代谢消耗的规律、起因与结果带来了挑战.本文首先介绍参与神经元编码的亚细胞过程及它们在大脑和小脑皮层中的代谢消耗,然后详细梳理近年来关于动作电位代谢消耗的研究成果,重点讨论影响其能量效率的生物物理因素和放电形状特性,并归纳总结放电消耗的特点,最后对未来神经元编码的代谢消耗研究进行展望.     

HCV复合多表位抗原基因表达及免疫原性的研究

分类号Ｑ７８１文献标识码Ａ文章编号０００１６２０９（１９９９）０３０２６８７１丙型肝炎是常见的传染病之一,易于慢性化及发展为肝硬化,且与肝癌的发生关系密切,目前尚无理想的防治手段。国内外对丙型肝炎疫苗的研究仍处于克隆表达ＨＣＶ部分基因产物或合成…     

Telemetry System for Assessing Jaw-Muscle Function in Free-ranging Primates

In vivo laboratory-based studies describing jaw-muscle activity and mandibular bone strain during mastication provide the empirical basis for most evolutionary hypotheses linking primate masticatory apparatus form to diet. However, the laboratory data pose a potential problem for testing predictions of these hypotheses because estimates of masticatory function and performance recorded in the laboratory may lack the appropriate ecological context for understanding adaptation and evolution. For example, in laboratory studies researchers elicit rhythmic chewing using foods that may differ significantly from the diets of wild primates. Because the textural and mechanical properties of foods influence jaw-muscle activity and the resulting strains, chewing behaviors studied in the laboratory may not adequately reflect chewing behaviors of primates feeding in their natural habitats. To circumvent this limitation of laboratory-based studies of primate mastication, we developed a system for recording jaw-muscle electromyograms (EMGs) from free-ranging primates so that researchers can conduct studies of primate jaw-muscle function in vivo in the field. We used the system to record jaw-muscle EMGs from mantled howlers (Alouatta palliata) at Hacienda La Pacifica, Costa Rica. These are the first EMGs recorded from a noncaptive primate feeding in its natural habitat. Further refinements of the system will allow long-term EMG data collection so that researchers can correlate jaw-muscle function with food mechanical properties and behavioral observations. In addition to furthering understanding of primate feeding biology, our work will foster improved adaptive hypotheses explaining the evolution of primate jaw form.     

Evaluating Global Positioning System Telemetry Techniques for Estimating Cougar Predation Parameters

ABSTRACT Using clusters of locations obtained from Global Positioning System (GPS) telemetry collars to identify predation events may allow more efficient estimation of behavioral predation parameters for the study and management of large carnivore predator-prey systems. Applications of field- and model-based GPS telemetry cluster techniques, however, have met with mixed success. To further evaluate and refine these techniques for cougars (Puma concolor), we used data from visits to 1,735 GPS telemetry clusters, 637 of which were locations where cougars killed prey >8 kg in a multi-prey system in west-central Alberta. We tested 1) whether clusters were reliably created at kill locations, 2) the ability of logistic regression models to identify kill occurrence (prey >8 kg) and multinomial regression models to identify the prey species at a kill cluster, and 3) the duration of monitoring required to accurately estimate kill rate and prey composition. We found that GPS collars programmed to attempt location fixes every 3 hours consistently identified locations where prey >8 kg were handled, and cluster creation was robust to GPS location acquisition failures (poor collar fix success). The logistic regression model was capable of estimating cougar kill rate with a mean 5-fold cross validation error of <10%, provided the appropriate probability cutoff distinguishing kill clusters from non-kill clusters was selected. Logistic models also can be used to direct visits to clusters, reducing field efforts by as much as 25%, while still locating >95% of all kills. The multinomial model overpredicted occurrence of primary prey (deer) in the diet and underpredicted consumption of alternate prey (e.g., elk and moose) by as much as 100%. We conclude that a purely model-based approach should be used cautiously and that field visitation is required to obtain reliable information on species, sex, age, or condition of prey. Ultimately, we recommend a combined approach that involves using models to direct field visitation when estimating behavioral predation parameters. Regardless of the monitoring approach, long continuous monitoring periods (i.e., >100 days of a 180-day period) were necessary to reduce bias and imprecision in kill rate and prey composition estimates.     

Assessment of Elk Highway Permeability by Using Global Positioning System Telemetry

Abstract: Highways have significant direct and indirect impact on natural ecosystems, including wildlife barrier and fragmentation effects, resulting in diminished habitat connectivity and highway permeability. We used Global Positioning System (GPS) telemetry to assess Rocky Mountain elk (Cervus elaphus nelsoni) permeability across a 30-km stretch of highway in central Arizona, USA, currently being reconstructed with 11 wildlife underpasses, 6 bridges, and associated ungulate-proof fencing. The highway was reconstructed in phases, allowing for comparison of highway crossing and passage rates during various stages of reconstruction. We instrumented 33 elk (25 F, 8 M) with GPS receiver collars May 2002 to April 2004. Our collars accrued 101,506 GPS fixes with 45% occurring within 1 km of the highway. Nearly 2 times the proportion of fixes occurred within 1 km of the highway compared with random. We think elk were attracted to the highway corridor by riparian—meadow foraging habitats that were 7 times more concentrated within the 1-km zone around the highway compared with the mean proportion within elk use areas encompassing all GPS fixes. Elk crossed the highway 3,057 times; crossing frequency and distribution along the highway were aggregated compared with random. Crossing frequency within 0.16-km highway segments was negatively associated with the distance to riparian—meadow habitats (r_s = -0.714, n = 190, P < 0.001). Mean observed crossing frequency (92.6 ± 23.5 [SE] crossings/elk) was lower than random (149.6 ± 27.6 crossings/elk). Females crossed 4.5 times as frequently as males. Highway permeability among reconstruction classes was assessed using passage rates (ratio of highway crossings to approaches); our overall mean passage rate was 0.67 ± 0.08 crossings per approach. The mean passage rate for elk crossing the highway section where reconstruction was completed (0.43 ± 0.15 crossings/approach) was half that of sections under reconstruction and control sections combined (0.86 ± 0.09 crossings/approach). Permeability was jointly influenced by the size of the widened highway and associated vehicular traffic on all lanes. Crossing frequency was used to delineate where ungulate-proof fencing yielded maximum benefit in intercepting and funneling crossing elk toward underpasses, promoting highway safety. Use of passage rates provides a quantitative measure to assess permeability, conduct future pre- and postconstruction comparisons, and to develop mitigation strategies to minimize highway impacts to wildlife.     

Component Object Based Single System Image for Dependable Implementation of Genetic Programming on Clusters

We present a distributed component-object model (DCOM) based single system image (SSI) for dependable parallel implementation of genetic programming (DPIGP). DPIGP is aimed to significantly and reliably improve the computational performance of genetic programming (GP) exploiting the inherent parallelism in GP among the evaluation of individuals. It runs on cost-effective clusters of commodity, non-dedicated, heterogeneous workstations or PCs. Developed SSI represents the pool of heterogeneous workstations as a single, unified virtual resource – a metacomputer, and addresses the issues of locating and allocating the physical resources, communicating between the entities of DPIGP, scheduling and load balancing. In addition, addressing the issue of fault tolerance, SSI allows for building a highly available metacomputer in which the cases of workstation failure result only in a corresponding partial degradation of the overall performance characteristics of DPIGP. Adopting DCOM as a communicating paradigm offers the benefits of software platform- and network protocol neutrality of proposed approach; and the generic support for the issues of locating, allocating and security of the distributed entities of DPIGP.     

Telemetry and Satellite Tracking of Whale Sharks, Rhincodon Typus, in the Sea of Cortez, Mexico, and the North Pacific Ocean

We used satellite-linked radio telemetry to document the geographic and vertical movements and thermal habitats of whale sharks in the Sea of Cortez and as they migrated into the north Pacific Ocean. Of 17 sharks tagged between 1994 and 1996, six dispersed widely in the Sea of Cortez during 12–39 days of tracking. Four others left the Sea of Cortez and ranged extensively in the north Pacific Ocean. Indeed, one whale shark migrated to the western north Pacific Ocean, covering over 13000km in 37 months of tracking. The sharks generally occupied areas where sea surface water temperatures were between 28 and 32°C, though several ranged to depths of 240m or deeper where water temperature reached 10°C or colder. Whale sharks may segregate by size and sex, and their movement patterns appear to be related to oceanographic features, such as sea mounts and boundary currents, where primary productivity may be enhanced. These results have important implications for the global conservation of the world's largest yet least known fish. We think that satellite telemetry is a exceptionally promising tool for learning more about the ecology of whale sharks, especially when combined with conventional methods of telemetry and molecular biology.authorship arranged alphabetically     

Florida Panther Habitat Selection Analysis of Concurrent GPS and VHF Telemetry Data

Abstract: Florida panthers (Puma concolor coryi) are listed as an endangered subspecies in the United States and they exist in a single Florida population with <100 individuals; all known reproduction occurs south of Lake Okeechobee. Habitat loss is the biggest threat to this small population and previous studies of habitat selection have relied on very high frequency (VHF) telemetry data collected almost exclusively during diurnal periods. We investigated habitat selection of 12 panthers in the northern portion of the breeding range using 1) Global Positioning System (GPS) telemetry data collected during nocturnal and diurnal periods and 2) VHF telemetry data collected only during diurnal periods. Analysis of both types of telemetry data yielded similar results as panthers selected upland (P < 0.001) and wetland (P < 0.001) forested habitat types. Our results indicated that forests are the habitats selected by panthers and generally support the current United States Fish and Wildlife Service panther habitat ranking system. We suggest that future studies with greater numbers of panthers should investigate panther habitat selection using GPS telemetry data collected throughout the range of the Florida panther and with location attempts scheduled more evenly across the diel period. Global Positioning System radiocollars were effective at obtaining previously unavailable nocturnal telemetry data on panthers; however, we recommend that panther researchers continue to collect VHF telemetry data until acquisition rates and durability of GPS collars improve. (JOURNAL OF WILDLIFE MANAGEMENT 72(3):633–639; 2008)     

Marine Telemetry and the Conservation and Management of Risk to Seal Species in Canada and Australia

Marine telemetry expands the knowledge of the biology of marine species at risk: their life cycles, activities, interactions, habitats, and threats. Four seal species in Canada and Australia are faced with distinctive and divergent management problems. This article examines their conservation status, legal protection, and the role that telemetry has played, or could play, in providing previously unavailable information to help meet conservation goals. The value of telemetry data to minimize fisheries mortality of one species has been demonstrated in Australia. Despite there being significant telemetry data for the other species, policy and management have not yet responded.     

Telemetry and genetics reveal asymmetric dispersal of a lake‐feeding salmonid between inflow and outflow spawning streams at a microgeographic scale

The degree of natal philopatry relative to natal dispersal in animal populations has important demographic and genetic consequences and often varies substantially within species. In salmonid fishes, lakes have been shown to have a strong influence on dispersal and gene flow within catchments; for example, populations spawning in inflow streams are often reproductively isolated and genetically distinct from those spawning in relatively distant outflow streams. Less is known, however, regarding the level of philopatry and genetic differentiation occurring at microgeographic scales, for example, where inflow and outflow streams are separated by very small expanses of lake habitat. Here, we investigated the interplay between genetic differentiation and fine‐scale spawning movements of brown trout between their lake‐feeding habitat and two spawning streams (one inflow, one outflow, separated by <100 m of lake habitat). Most (69.2%) of the lake‐tagged trout subsequently detected during the spawning period were recorded in just one of the two streams, consistent with natal fidelity, while the remainder were detected in both streams, creating an opportunity for these individuals to spawn in both natal and non‐natal streams. The latter behavior was supported by genetic sibship analysis, which revealed several half‐sibling dyads containing one individual that was sampled as a fry in the outflow and another that was sampled as fry in the inflow. Genetic clustering analyses in conjunction with telemetry data suggested that asymmetrical dispersal patterns were occurring, with natal fidelity being more common among individuals originating from the outflow than the inflow stream. This was corroborated by Bayesian analysis of gene flow, which indicated significantly higher rates of gene flow from the inflow into the outflow than vice versa. Collectively, these results reveal how a combination of telemetry and genetics can identify distinct reproductive behaviors and associated asymmetries in natal dispersal that produce subtle, but nonetheless biologically relevant, population structuring at microgeographic scales.     

Validation of a New Video and Telemetry System for Remotely Monitoring Wildlife

Abstract: Current techniques for remotely monitoring wildlife lack the capability to survey a wide area or to transmit data in real time. We addressed these technology gaps by developing and testing a new video and telemetry system for remotely sampling wildlife abundance, distribution, and behavior across large open areas. The system consisted of 2 pan-tilt-zoom video cameras equipped with 20–200× lens, and an automated telemetry scanner and data logger. All components were charged by wind and solar power and located on a hilltop overlooking an open valley (23 km²) in Yellowstone National Park, USA. A satellite up-link to the internet transmitted data in real time to the University of Minnesota-St. Paul and relayed commands from undergraduate students who controlled the cameras and systematically scanned the area at 2-hour intervals 6 times/day (0800–2000 hr) 7 days/week for about 20 consecutive weeks (Dec-Jun). During each scan, students recorded presence, activity, and location of bison (Bison bison), coyote (Canis latrans), grizzly bear (Ursus arctos), red fox (Vulpes vulpes), and wolf (Canis lupus). The telemetry system continuously scanned frequencies of 100 tagged wolves, including 4 members of the one resident pack. We determined wolf presence in real time by viewing the incoming data stream, or we assessed presence later after off-loading the data logger. Matched pairs of simultaneous observations taken by remote and on-site observers during a 13-day double-sampling period were highly correlated (r = 0.71–0.94), but remote observations were biased toward larger, more visible mammals (e.g., bison) and tended to underestimate their abundance. Nevertheless, the system was deployed longer than was practical for on-site observers and was, therefore, useful for detecting long-term, fine-scale trends such as daily changes in bison numbers as a function of snow depth. (JOURNAL OF WILDLIFE MANAGEMENT 72(8):1834–1844; 2008)     

Development of an In vitro pod culture technique for young pods of Phaseolus vulgaris L.

Summary A research program to develop an in vitro culture technique adapted to very immature Phaseolus zygotic embryos indicated that osmolality within young pods in vivo ranged from 580 to 350 mosm during the first 10 d after pollination. Different culture techniques for 2-d-old Phaseolus pods are described using a modified Phillips medium for maturation. The application of high and variable osmolality conditions, similar to what is observed in vivo, during pod culture (1 wk) and before extracting the embryos, gave the best results in terms of ovule and embryo development. To solve contamination problems affecting pod development during these investigations, a sterilizing technique was tested combining various concentrations of polyoxyethylenesorbitan monolaurate (Tween 20) and a broad-spectrum industrial biocide, Plant Preservative Mixture (PPM). The PPM sterilizing method totally eliminated pod contamination and did not affect ovule growth, but significantly reduced the embryo germination rate.