本学期开始，我要给本科生上《现代生物科学导论》这门课了。虽然读大学的时候系统学习过与生物学相关的一些课程，但20多年过去了，变化很大，看到教材，发现许多内容对我来说仍然是相当陌生的。因此，如果能有机会向一些有授课经验的老师请教，那将是非常好的事。上周日，学院提供了这样一个机会，我当然求之不得，第一个到了会场等候着。的确，从有经验的授课老师那里，我学到了不少东西。 教案分析中，有老师提供了跳蛛（zebra spider）与雪蝶（snowberry fly）的经典拟态故事：与其他蛛类结网捕食不同，跳蛛是跟踪捕猎，在猎物后面蹑手蹑脚地，然后逮着机会猛扑过去。跳蛛的领地意识非常浓厚，它们会极力保卫自己的领地，当其他同类来犯时，它们会挥舞着腿以示抗议，直到入侵者逃跑。雪蝶是跳蛛喜欢的食物。雪蝶的透明翅膀上有一些黑色标记，看起来非常像跳蛛的腿。当雪蝶碰到跳蛛时，就会舞动翅膀，这些标记和行为似乎就像跳蛛挥舞着腿一样，这种模仿捕食者的拟态行为是不多见的，但的确是生态学中非常有趣的范例（ 详细内容，请参见网站 ）。 不过，为了验证这样的行为是否属实，就有人设计了实验，将雪蝶的翅膀全部涂成黑色作为处理，与不涂翅膀的雪蝶进行对比，看最后被跳蛛所捕食的效果。实验显然是成功的，翅膀被涂成黑色的雪蝶被捕食的几率要大得多。这个实验看似没有问题吧，但是严格地说，这个实验并不完善：将雪蝶的翅膀涂黑这个操作中，不能假设雪蝶的翅膀没有受到其他的损害（比如划伤，或者变得湿润）。因此，这个看似单一的处理，其实是两个处理（涂色+对翅膀的潜在损伤）。所以，要完善这个实验，必须另加一个处理：用透明的染料将翅膀涂一遍。如果最终这一组处理与对照组没有显著差异，而与涂黑组有显著性差异，这个结果才算有效。 跳蛛与雪蝶 被跳蛛所捕获的雪蝶所占的百分比 由于早上讨论了这个问题，晚上看巴西科学家的这篇论文的时候，很快就联想到了实验设计的严密性问题。实话说，我不知道，Bt微生物制剂究竟是什么样的（如果知道的，请帮忙给一个权威的说法）。从一些网页的描述中，我理解这个制剂所含的微生物是没有灭活的，也就是含菌株的。例如，有些网站是这样描述的：“ 由于微生物可以被雨水冲洗掉，且在喷洒农药后的几天内Cry蛋白会在阳光下失去活性，因而Bt微生物施用于叶子表面进行杀虫的效果有限 ” 。我理解巴西的这个实验，就是用Bt微生物制剂（农业中广泛施用的那种微生物农药）作为一个处理来进行的，其对照就是蒸馏水。由于Bt微生物制剂是溶于37°C环境的蒸馏水中的，这样的温度，如果含菌株，细菌开始活动是无疑的。我开始怀疑这个实验设计是否存在不合理的问题：单一的处理中可能含有多个复合因子，比如微生物本身所产生的影响，我认为需要增加一个处理——含微生物但没有分泌孢子晶体。想到这里，我觉得应该写篇博文来表达一下自己的认识。另外，由于之前读到蒋高明博主的博文，将这个实验与转基因食品的安全风险联系起来，我就有了一个先入为主的认识（后来，我又认真看了一遍论文，发现论文的内容与转基因食物没有什么关系）。我在博文“ 食用转基因食品会导致白血病的说法，靠谱吗？ ”中，对这个疑问进行了论述： 这项研究根本就没有给小鼠喂食任何转基因食品，只是给小鼠喂食了含苏云金杆菌的悬浊液而已！也就是说，从这个实验得出的结果，既与转基因食品无关，也与Bt蛋白无关，充其量只能说明这些血液方面的效应是来自苏云金杆菌的，苏云金杆菌并不能与Bt蛋白划等号。 也就是说，从这个实验得出的结果，与转基因食品并无任何关联。这些论述，试图从实验设计的角度提示一下可能对科学实验进行捕风捉影的衍生。 换一个角度来看，巴西科学家的这篇论文，更像是对反转者的高级黑。所以我在前面的那篇博文（修改版）中这样论述的： 就这篇论文的研究来说，能让我们直接联想到的，并不是转基因作物，而是绿色食品、有机食品中广泛使用的含Bt微生物制剂。如果质疑这个微生物制剂的安全性得到证实，首先受到打击的应该是这些绿色有机食品的生产，其次才可能是转基因作物，或者二者同时受到了挑战。 另外一个问题，也是从实验设计的角度来说的。设计一个毒理学实验，什么剂量才算是合适的？巴西的实验中，分别采用了27mg/Kg, 136mg/Kg和270mg/Kg这几个水平，原文是这样描述的： The minimum dose of Bt sporecrystal toxins was of 27 mg/Kg; the maximum dose (270 mg/Kg) was 10 times greater than the minimum dose, while the intermediate dose (136 mg/Kg) was equivalent to about half the maximum dose 。 隔行如隔山，我对这个剂量并没有什么感觉，按照巴西论文中的说法： In our previous experiments, exposures greater than 270 mg/Kg had caused signs of toxicity and death, so this concentration was considered the maximum tolerated. 也就是说，大于270mg/Kg可能是致死剂量了，我理解这是很大的量。后来看到一个英文网站 （ http://www.biofortified.org/2013/05/leukemia/ ） 中对这个剂量的评价： The amounts fed the mice do not reflect human dietary levels: they were some ~ 10^6 to 10^8 time higher than exposure from GM or organic crops (as per Hammond, 2012)。也就是说，是人类实际在转基因作物和有机作物下能暴露剂量的1百万到1亿倍（这个数值有待行家论证）。所以，我在博文中这样论述： 任何化学药品，剂量增加到一定程度都会构成危害，因此这样的结果意义不大。 比如，换用食盐做类似的实验：用致死剂量下的几个梯度进行，观测血液方面的指标，相信也会有许多的变化。但这样的变化，可解释性并不强。 我只是用自己的认识和常识来质疑巴西这篇文章的方法和推理方面的问题，与转基因作物问题相比，我更在乎其实验设计的合理性。也许有局部误解的地方。这样的想法放在自己的博客空间，其实很正常的事儿了（科学网编辑推荐，那是他们的事儿，与我无关），大家一起讨论，就是为了将问题弄清楚。在转基因问题上有明显立场的人，似乎无法容忍任何与自己观点不同的看法，总是喜欢搞一些攻击，这一点很容易打破客观讨论的气氛（所有与自己观点不一致的人都是造谣吗？）。我从来没有标榜自己是转基因研究方面的专家，但也不是一无所知。
名称 Experimental Design and Data Analysis for Biologists 作者 Gerry P. Quinn Monash University, Victoria Michael J. Keough University of Melbourne An essential textbook for any student or researcher in biology needing to design experiments, sample programs or analyse the resulting data. The text begins with a revision of estimation and hypothesis testing methods, covering both classical and Bayesian philosophies, before advancing to the analysis of linear and generalized linear models. Topics covered include linear and logistic regression, simple and complex ANOVA models (for factorial, nested, block, split-plot and repeated measures and covariance designs), and log-linear models. Multivariate techniques, including classification and ordination, are then introduced. Special emphasis is placed on checking assumptions, exploratory data analysis and presentation of results. The main analyses are illustrated with many examples from published papers and there is an extensive reference list to both the statistical and biological literature. The book is supported by a web-site that provides all data sets, questions for each chapter and links to software. An essential textbook that covers the full range of topics required by any biologist designing experiments or analysing the resulting data Extensive worked examples using real data sets Web page support with downloadable data sets, example questions and relevant links at http://www.zoology.unimelb.edu.au/qkstats Contents 1. Introduction; 2. Estimation; 3. Hypothesis testing; 4. Graphical exploration of data; 5. Correlation and regression; 6. Multiple regression and correlation; 7. Design and power analysis; 8. Comparing groups or treatments - analysis of variance; 9. Multifactor analysis of variance; 10. Randomized blocks and simple repeated measures: unreplicated two-factor designs; 11. Split plot and repeated measures designs: partly nested anovas; 12. Analysis of covariance; 13. Generalized linear models and logistic regression; 14. Analyzing frequencies; 15. Introduction to multivariate analyses; 16. Multivariate analysis of variance and discriminant analysis; 17. Principal components and correspondence analysis; 18. Multidimensional scaling and cluster analysis; 19. Presentation of results. Reviews At last, a book that provides a readable introduction to nuances of statistical methods and analysis a wonderful book that is packed with lots of practical advice . Journal of Experimental Marine Biology and Ecology this is clearly written text with a simple no-nonsense approach to the topic. TEG News ' the book is well written and well presented with a good range of interesting and realistic examples the book gave a very substantial and worthwhile study of good statistical practice in the design and analysis of biological experiments. I recommend it to anyone involved in quantitative biological research.' Journal of Agricultural Science Quinn and Keough make plenty of reference to the recent and primary statistical literature, yet their book does not seem inaccessible or daunting the text often ventures into statically uncertain territory, and Quinn and Keough do an excellent job of evenhandedly summarizing any statistical debates and philosophies then giving pragmatic suggestions to how best to proceed with analyses. Readers will find themselves adequately and interestedly informed Quinn and Keough make extensive use of data sets deriving from real, and recently published, studies There are also unexpected bonus sections, such as the useful, and at times fun, chapter on presenting the results of analysis both in reports and in seminars. In general, one certainly has the impression that the authors set out to write a clear, comprehensive and valuable book: they have succeeded.' Animal Behaviour highly recommended Ethology ' the authors do go a long way towards success in their aim of encouraging 'readers to understand the models underlying the most common experimental designs' and to approach proper data analysis with more confidence. The web support is also very useful especially for items that the authors added post-publication '. Primate Eye an essential textbook that can be warmly recommended to any student or researcher in biology who needs to design experiments, devise sampling programs and analyze the resulting data There is a wealth of information that is usually only found in separate sources. Basic and Applied Ecology ' an essential textbook for students and researchers in biology needing to design experiments, sampling programs or analyze the resulting data.' Folia Geobotanica
标签: 实验设计
