非调制式荧光仪PEA测定叶绿素荧光参数的研究

为了利用非调制式荧光仪获得调制式荧光仪测定的叶绿素荧光动力学参数,采用植物效率(PEA)仪进行了叶绿素荧光参数测定程式的4要素(光化光强度及其照射时间、饱和激发光强度及其照射时间)以及节约测定时间和仪器内存的测定技术研究。结果表明,暗适应10min后,在设定180μmol·m-2·s-1光化光照射360s时间内,以连续2次1950μmol·m-2·s-1饱和激发光照射3s的测定程式,在7种植物的叶绿素荧光参数的测定中获得了较为满意的测定结果,而且与Strasser等测定程式相比,1次测定需时从600s缩短为360s、耗用仪器内存从76.9"减少为2.56"。     

青海高原及上海平原地区植物叶片光合作用的光抑制

用便携式ＡＤＣ光合气体分析系统和便携式ＣＦ－１０００荧光仪对青海和上海的同一植物和不同植物叶片光合作用的光抑制进行了测定。结果表明,两地在晴天强光下,中午植物叶片光系统Ⅱ（ＰＳⅡ）的光化学效率（Ｆｖ／Ｆｍ）和表观光合量子效率（ＡＱＹ）的日变化比早晨低;上海测定ＰＳⅡ的Ｆｖ／Ｆｍ和ＡＱＹ的日变化比青海的下降幅度大。ＡＱＹ的日变化曲线比ＰＳⅡ的Ｆｖ／Ｆｍ低,ＡＱＹ降低的幅度比ＰＳⅡ的Ｆｖ／Ｆｍ大。两地植物均有不同程度的光合作用光抑制,且上海比青海的大     

濒危植物长序榆(Ulmus elongata)幼苗叶绿素荧光特性的初步研究

用Li-6400XT便携式光合作用仪对濒危植物长序榆幼苗的各叶绿素荧光参数的日变化和快速光响应曲线进行了测定。结果发现,光系统Ⅱ(PSⅡ)的实际光化学效率(ΦPSⅡ)、电子传递速率(ETR)在整个白天阶段较稳定,下午18:00显著下降。光化学淬灭(qP)先增大后减小。非光化学淬灭(NPQ)呈现出与光化学淬灭(qP)相反的变化趋势,中午最低,说明长序榆幼苗光能利用率较高。快速光曲线表明实际光化学效率(ΦPSⅡ)和光化学淬灭(qP)随着光合有效辐射(PAR)的增大而减小,电子传递速率(ETR)和非光化学淬灭(NPQ)随着光合有效辐射(PAR)的增大而增大。使用幂函数能够很好的拟合实际光化学效率(ΦPSⅡ)和电子传递速率(ETR)随光强的变化,而对数函数能较好的拟合实际光化学淬灭(qP)和非光化学淬灭(NPQ)随光强的变化。     

用体内叶绿素a荧光诱导动力学鉴定番茄的抗冷性

研究了冷害温度对具有不同抗冷性品种的番茄叶片的体内叶绿素ａ荧光诱导动力曲线的影响。实验结果指出,在低温处理（８℃,５℃,２℃下,暗中２４小时）后,番茄叶片的体内叶绿素ａ荧光诱导动力学曲线有了明显的改变,Ｆｖ／Ｆｏ值、Ｒｆｄ值降低了,光系统Ⅱ原初光能转换效率和潜在的光合活力均受到抑制。我们在苗期和开花期得到的实验结果均表明,在番茄叶片的叶绿素ａ荧光诱导动力学曲线和这些荧光参数改变的程度与该品种的已知抗冷性之间呈现较好的相关性。我们认为,体内叶绿素ａ荧光诱导动力学方法是鉴定番茄抗冷性的一个快速、灵敏和可靠的方法,并可用于其他绿色植物的抗冷性鉴定中     

蝴蝶兰黄花品系光合能力快速测定

以5个蝴蝶兰黄花品系为试材,利用调制叶绿素荧光技术,通过比较其快速光曲线相关参数,以了解不同品系光合能力,尝试建立蝴蝶兰光合能力快速测定技术。结果表明：不同蝴蝶兰黄花品系光合能力存在明显差异,具有较高半饱和光强的品系也具有较高的最大相对电子传递速率,而初始斜率在一定程度上决定了品系最大相对电子传递速率的高低。调制叶绿素荧光技术不仅可用于蝴蝶兰光合能力快速测定,还可应用于蝴蝶兰其他光合生理研究。     

植物ms级叶绿素荧光动力学数据采集和分析软件

本文介绍了一套可在ＩＢＭＰＣ（兼容）微机上运行的植物ｍｓ级荧光动力学数据采集和分析软件。本软件用Ｃ语言写成,具有汉化界面、菜单驱动、彩色人机对话窗口、操作简便、灵活,兼容性好等优点。本文着重讨论了本软件如何减少荧光动力学测量中的误差、在不过分占用内存的情况下扩大采样时间范围以及如何精确测量固定荧光（Ｆｏ）和偏转荧光（Ｆｉ）等问题。本软件和我们自行组装的植物ｍｓ级动力学荧光计———植物产量荧光计ＰＦＭ１０１型可广泛应用于农业、植物生态学及植物生理学的光合测量和研究     

高温胁迫下苋菜的叶绿素荧光特性

为了探明高温胁迫对苋菜(Amaranthus tricolor L.)光合过程的影响,用不同温度(25、30、35、40、45℃)处理苋菜植株1h后,随即测定了其叶绿素荧光动力学参数和快速光响应曲线特征参数的变化.结果表明:40℃以上高温胁迫下,苋菜叶片的光系统Ⅱ(PSⅡ)潜在光化学效率(Fv/Fo)、最大光化学效率(Fv/Fm)下降;最大荧光(Fm)、光合电子传递速率(ETR)、PSⅡ实际光化学效率(Yield)、光化学淬灭系数(qP)也均有所下降;而初始荧光(F.)和非光化学淬灭系数(NPQ)在40℃以上高温胁迫下显著上升.叶绿素荧光快速光响应曲线测定结果表明,初始斜率α、最大相对电子传递速率ETRmax和半饱和光强Ik在40℃以上高温胁迫下有所下降.研究表明,40℃以上高温胁迫对苋菜的光能的吸收、转换、光合电子传递和强光耐受能力等均有一定的影响.     

几种常绿植物光合特性的季节变化

以在上海自然条件下生长的瑚瑚树、女贞、孝顺竹和棕榈等四种常绿植物为材料,在春、夏、秋、冬四个季节测定了它们的叶片在不同温度时的光饱和光合速率;用ＣＦ１０００荧光仪测定了叶片的初始光（Ｆｏ）、最大荧光（Ｆｍ）和可变荧光（Ｆｖ）等特征值．结果表明．瑚瑚树和女贞适应环境温度变化的能力较强,它们的光合最适温度（Ｔｏｐｔ）的变化范围比较大,在四季均保持较高的光合活力,光合量子效率和Ｆｖ／Ｆｍ值较高,而孝顺竹和棕榈适应环境温度变化的能力较差,只在春秋两季有较高的光合活力．这些差别和它们的自然分布有联系．     

水分胁迫对小麦旗叶某些体内叶绿素a荧光参数的影响

研究了水分胁迫对京冬６号品种小麦（ＴｒｉｔｉｃｕｍａｅｓｔｉｖｕｍＬ．）的旗叶（于杨花期）的体内（ｉｎｖｉｖｏ）叶绿素ａ荧光诱导动力学及几种叶绿素ａ荧光参数的影响,以探究水分胁迫损伤光合作用的机理。通过将离体叶片放置于干燥器中不同时间以得到三种干旱的情况（失水叶片的ＲＷＣ分别为５０．９％,３１．５％和１７．５％。对照叶片的ＲＷＣ为７３．５％）。体内叶绿素ａ荧光特性是用ＰＡＭ叶绿素荧光计测定的。我们观察到水分胁迫显著地改变了小麦旗叶的重复饱和光脉冲激发的调制叶绿素ａ荧光诱导动力学的图形。对从这种荧光动力学的实验数据得到的四种重要的体内叶绿素ａ荧光参数：Ｆｍ／Ｆｏ值（最大荧光与基础荧光的比值）、（ｇ－ｈ）值（饱和光脉冲诱导的荧光尖峰信号高度）、ｑＰ值（光化学淬灭系数）和ｑＮ值（非光化学淬灭系数）,水分胁迫都有抑低的作用。并且我们注意到,在水分胁迫的严重程度与这四种荧光参数被抑低的程度之间,存在着正相关关系。我们的实验数据指出,水分胁迫抑制了光系统Ⅱ的氧化侧,并且也降低了光合膜的能量化作用     

用体内叶绿素a荧光诱导动力学鉴定番茄的抗冷性

研究了冷害温度对具有不同抗冷性品种的番茄叶片的体内叶绿素a荧光诱导动力曲线的影响。实验结果指出,在低温处理(8℃,5℃,2℃下,暗中24小时)后,番茄叶片的体内叶绿素a荧光诱导动力学曲线有了明显的改变,Fv／Fo值、Rfd值降低了,光系统II原初光能转换效率和潜在的光合活力均受到抑制。我们在苗期和开花期得到的实验结果均表明,在番茄叶片的叶绿素a荧光诱导动力学曲线和这些荧光参数改变的程度与该品种的已知抗冷性之间呈现较好的相关性。我们认为,体内叶绿素a荧光诱导动力学方法是鉴定番茄抗冷性的一个快速、灵敏和可靠的方法,并可用于其他绿色植物的抗冷性鉴定中。     

植物ms 级叶绿素荧光动力学数据采集和分析软件

本文介绍了一套可在IBM PC(兼容)微机上运行的植物ms级荧光动力学数据采集和分析软件。本软件用C语言写成,具有汉化界面、菜单驱动、彩色人机对话窗口、操作简便、灵活,兼容性好等优点。本文着重讨论了本软件如何减少荧光动力学测量中的误差、在不过分占用内存的情况下扩大采样时间范围以及如何精确测量固定荧光(Fo)和偏转荧光(Fi)等问题。本软件和我们自行组装的植物ms级动力学荧光计——植物产量荧光计PFM-101型可广泛应用于农业、植物生态学及植物生理学的光合测量和研究。     

光片荧光显微镜特点及其应用

传统荧光显微镜由于对某些荧光分子存在光毒性、光损伤等方面的缺陷,无法满足对部分活体样本进行长时间观测的需求。光片荧光显微镜（light sheet fluorescence microscope,LSFM）是一种新型荧光显微镜,有别于激光共聚焦显微镜,其特殊的正交光路设计和高效的信号采集装置,使其具备低光毒性、低光漂白、低光损伤和高时空分辨率等优良特性,从而能对细胞及大尺度生物组织样本进行时空连续性较好的记录,尤其适宜于活体生物样品。基于此,概述了光片荧光显微镜的成像原理、成像优势、成像效果的改进与优化历程及其在生命科学领域应用所取得的研究成果,重点对近三年相关应用进行了汇总,并简要介绍了其在神经生物学、发育生物学、动物细胞生物学和植物科学领域中一部分代表性研究内容,最后,总结了光片荧光显微镜的优点与发展至今仍存在的不足,并对其在光遗传学和多组学研究中的潜在应用进行了展望,以期为研究人员提供较为系统的光片荧光显微镜相关基础知识、最新的研究应用进展以及未来的潜在应用方向,为研究人员提供参考。     

Growth of normal oral keratinocytes and squamous cell carcinoma cells in a novel protein-free defined medium

Summary A novel protein-free synthetic medium was developed for the culture of normal human oral keratinocytes. This medium, designated PFM-7, supports the serial cultivation of primary or secondary normal oral keratinocytes in protein-free, chemically defined conditions. Normal oral keratinocytes in PFM-7 exhibited nearly equal growth in mass culture without noticeable changes in morphology, response to added growth factors, or gene expression of growth factors and their receptors, compared to cells in Keratinocyte-SFM containing epidermal growth factor and bovine pituitary extract. Furthermore, PFM-7 supported the serial subcultivation of human squamous cell carcinoma cells and enabled both normal and malignant oral squamous cells derived from the same patient to grow under the same protein-free defined conditions. These results indicate that PFM-7 can be used for precise investigations of growth mechanisms, cell products, and gene expression associated with carcinogenesis of human epidermal cells.     

The Kautsky curve is a built-in barcode.

We identify objects from their visually observable morphological features. Automatic methods for identifying living objects are often needed in new technology, and these methods try to utilize shapes. When it comes to identifying plant species automatically, machine vision is difficult to implement because the shapes of different plants overlap and vary greatly because of different viewing angles in field conditions. In the present study we show that chlorophyll a fluorescence, emitted by plant leaves, carries information that can be used for the identification of plant species. Transient changes in fluorescence intensity when a light is turned on were parameterized and then subjected to a variety of pattern recognition procedures. A Self-Organizing Map constructed from the fluorescence signals was found to group the signals according to the phylogenetic origins of the plants. We then used three different methods of pattern recognition, of which the Bayesian Minimum Distance classifier is a parametric technique, whereas the Multilayer Perceptron neural network and k-Nearest Neighbor techniques are nonparametric. Of these techniques, the neural network turned out to be the most powerful one for identifying individual species or groups of species from their fluorescence transients. The excellent recognition accuracy, generally over 95%, allows us to speculate that the method can be further developed into an application in precision agriculture as a means of automatically identifying plant species in the field.     

圆锥南芥适应高山环境极端高温的系统性耐热能力

高山生态系统的主要气候特征是温度变化幅度较大,目前研究主要集中于高山植物的抗冻机制,而很少关注其对极端高温（高于45℃）的适应性。本研究发现高山物种圆锥南芥跟拟南芥比,具有更强的基础耐热性和获得性耐热性。通过叶绿素荧光检测发现在极端高温下,圆锥南芥具有更稳定的光系统II和更有效的能量耗散机制来维持更高水平的光合效率。通过电导率和丙二醛含量的检测发现圆锥南芥的膜伤害更小,膜流动性与脂肪酸链的长度和不饱和度紧密相关,圆锥南芥脂肪酸具有更低的16∶3含量,更长的碳链,不饱和度没有明显的变化,这些可能有助于维持膜的稳定。另外,更高表达量的HSP101和HSP70可能为圆锥南芥提供了更好的保护作用。以上结果表明,圆锥南芥能利用生理生化活动的调整来适应高山环境中的极端高温,这种耐热策略与低地耐热植物相似,因此圆锥南芥具有系统性耐热能力,可以作为研究植物耐热分子机制的模式物种。     

Adaptation of a PAM-fluorometer for remote sensing of chlorophyll fluorescence

The Laser-PAM described in this paper is an adaptation of the PAM 101 fluorometer (Heinz Walz, Effeltrich, Germany) designed for remote sensing and non-invasive laboratory measurements of chlorophyll fluorescence. It is based on a 5 mW laser diode, emitting at 638 nm, and a Fresnel lens coupled to the ED-101 detection unit. Due to these modifications, measurements can be performed at a distance ranging from 0.3 to 2 m. The ED-101 detection unit has been modified to perform simultaneous measurements of both modulated fluorescence and light reflected by the leaf. Reflected light showed a good estimation of the photosynthetically active radiation measured exactly at the same area as the fluorescence. A particular advantage of the Laser-PAM fluorometer is its suitability for remote measurements under field conditions. Simultaneous fluorescence and gas-exchange measurements, performed on grapevine leaves, are reported as an example of an application for the Laser-PAM. This revised version was published online in June 2006 with corrections to the Cover Date.     

Use of the fluorescent timer DsRED-E5 as reporter to monitor dynamics of gene activity in plants

Fluorescent proteins, such as green fluorescent protein and red fluorescent protein (DsRED), have become frequently used reporters in plant biology. However, their potential to monitor dynamic gene regulation is limited by their high stability. The recently made DsRED-E5 variant overcame this problem. DsRED-E5 changes its emission spectrum over time from green to red in a concentration independent manner. Therefore, the green to red fluorescence ratio indicates the age of the protein and can be used as a fluorescent timer to monitor dynamics of gene expression. Here, we analyzed the potential of DsRED-E5 as reporter in plant cells. We showed that in cowpea (Vigna unguiculata) mesophyll protoplasts, DsRED-E5 changes its fluorescence in a way similar to animal cells. Moreover, the timing of this shift is suitable to study developmental processes in plants. To test whether DsRed-E5 can be used to monitor gene regulation in plant organs, we placed DsRED-E5 under the control of promoters that are either up- or down-regulated (MtACT4 and LeEXT1 promoters) or constitutively expressed (MtACT2 promoter) during root hair development in Medicago truncatula. Analysis of the fluorescence ratios clearly provided more accurate insight into the timing of promoter activity.     

Transport of Fluorescence through Highly Scattering Media: Corrections to the Determination of Quantum Yields

The equations for transport of fluorescence, generated within a highly scattering medium, are solved within the boundary conditions of the Kubelka and Munk treatment. Expressions are derived in closed form for fluorescence fluxes emanating from the front and back surfaces of a highly scattering infinite slab, whether or not fluorescence is absorbed within the sample. An “apparent” quantum yield, calculated from observed intensities of fluorescence and of back-scattered light from the front surface of the slab, can be corrected by these expressions to return the true quantum yield of fluorescence. Allowance for re-emission of reabsorbed fluorescence can be made, but may not be applicable in some experimental arrangements. Calculations performed on the fluorescence of rhodamine 101 suggest that in typical practical situations the correction factor may not be far from unity.