一些生醫研究的迷思 A few paradox in biomedical research

一些生醫研究的迷思A few paradox in biomedical research 賴明德 Ming-Derg Lai Department of Biochemistry and Molecular Biology College of Medicine National Cheng Kung University

If at first, the idea is not absurd, then there is no hope for it --A. Einstein You are here!

Dilemma and Choices for PhD Student • Find an advisor • Choose the research topic • Perform the experiments • Write a paper or proposal • What seminars do I need to attend? • Do I need to become a Lab’s favorite?

選擇那一個研究領域？個人興趣或熱門領領域？選擇那一個研究領域？個人興趣或熱門領領域？ • 科學研究成功第一要素：熱情enthusiasm • 你可能對某些領域有強烈之喜好或排斥，但背後形成的原因卻經常是教師所表現的技巧或熱心，而不是你真正的興趣，或是特定主題所具備的重要性。Enthusiasm is infectious. • 學生們常以自己以前作的不錯之題目，當成日後研究領域之選擇基準。 • 事實上主題範圍最適合用[重要性]判斷，不宜太設限在狹隘之幾個領域。

有效率或有創意的實驗室？ • We may have many more papers published when we work in a laboratory performing research efficiently. • We may learn independence and perhaps be more creative in a less-organized laboratory. • 李遠哲作為一個例子。

作可畢業或可能有大突破之題目？ • 創造自我科學研究之意義或按部就班的循序漸進達到成功？創造自我科學研究之意義經常導致失敗對有修業年限之博士班學生可能不宜？ • 有突破性之題目並非經常可遇到 • 重要性為第一考量，對一個重要題目之支節小題目先進行研究。把研究提案放在一個〝大場景〞去看 • Again, your enthusiasm is the most important!

研究的意義 • The most immediate benefit is going to be doing experiments that teach us things • Andrew Z. Fire, 2006 Nobel Laureate

研究的意義 • Part of the pleasure and the fascination of what we do has been in discovering the mechanics, the inner workings of that machine. • Roger D. Kornberg • The Nobel Prize in Chemistry 2006

假說型或非假說型研究？ • 一般而言，假說型實驗較受科學家喜歡。 • 假說型實驗之決定點在於〝新穎性〞及〝可行性〞。其危險不穩定性隨新穎性增加而增高。

根據最近Nature paper data產生之想法或根據實驗室之數據產生想法？ • The new idea is better based on the (1) preliminary experimental finding in our own lab (2) presentation in a seminar with unpublished result (3) data in a prestigious journal, but not in a well-known journal, such as Nature and Science. • Assimilate the new idea from the novel paper in the original research direction.

Well, we were led to it pretty much by our experimental noses. The people in the plant field had done tremendous work on gene silencing and so we, sort of, were following in their footsteps in trying to sort out what was responsible for this weird silencing phenomenon in the worm. • Andrew Z. Fire

Hypothesis versus Descriptive • It would be wrong to make too much of discoveries. The greatest advances in molecular biology have grown out of the intent and unrelaxing study of characteristics of a single biological system, E. coli. • Other examples: (1) Knockout mice experiments lead to important scientific developments. (2) Structural study or structure-function study are mostly non-hypothesis based. (3) Assay of MAPK, JNK, ERK pathways, systematic analysis • 我沒有偉大夢想，只有腳踏實地賣力工作 —Anil D. Ambanl

Weak rationale experiment • Advisor always asks me to try this or that. It seems that he/she does not have a clear aim and does not know where research goes. • 只有當投資人不曉得自己在幹什麼時，才必須分散投資—Warrin Buffett • This is an indication of non-hypothesis experiment or weak-rationale experiment. • Novel breakthrough frequently comes from weak-rationale experiments! • Why don’t you propose your own hypothesis and prove the correctness of this “novel” hypothesis in such occasions?

Research Topics • Unrelated looking stories just keep coming together and unfolding in beautiful ways • Craig C. Mello, The Nobel Prize in Physiology or Medicine 2006

Design an Experiment • 試用畫圖解表示問題。 • 將問題以明確陳述句表示。 • No experiments should be undertaken without a clear preconception of the forms its result might take. (you will include adequate control and have well-designed experiment by this way)

How to design a positive control and negative control • Write the question you ask (將問題以明確陳述句表示)。 • HBV L protein directly interact with ApoH • 改變名詞中的變項，使形成control. • P53 protein directly interact with Large T antigen (positive control) • HBV L protein directly interact with Large T antigen (negative control) • p53 protein directly interact with Apo H (negative control)

Design an experiment: How many controls do I need ? • Internal control is absolutely required. Actin in western control, GAPDH in RT-PCR. • Negative control is required. • To investigate whether AKT is required for c-fos induction. Cells cotransfected with AKT siRNA and c-fos luciferase reporter, the luciferase is measured. pCMV-galactosidase is used as internal control. The scrambled Rnai is used as negative control. • Positive control is highly recommended when the result is positive. Positive control is required when the result is negative. • Cells treated with tunicamycin (ER stress inducer), and KCC4 ion transporter is measured. The GRP78 as a positive control for the induction of ER stress.

Start a pilot experiment: How many variables can be tested? • Based on the tolerable marginal error (time and money consumption) to determine (1) how many variables will be tested. (2) how much you can trust authority. • Test the reasonable variable by N + (N-1) rule,test one variable at a time. Nat Rev Mol Cell Biol. 2004 Jul;5(7):577-81

作實驗應嚴謹follow protocol 或進一步改良實驗方法 • 先了解protocol 之chemical or physical principle. • 記錄修改protocol 之地方 • 實驗成功後，不得修改protocol when repeating experiments!

嚴謹遵循實驗流程卻無法得到結果？手藝不靈巧？嚴謹遵循實驗流程卻無法得到結果？手藝不靈巧？ • 生醫實驗尚有許多因子無法精確掌握，for example, the times of subculture and the cell density at subculture will affect the experimental result. • 控制實驗細節 • 進行模擬實驗(virtual experiment): Do a mental dry run, to be sure that you have not forgotten anything. • 分割實驗 • 減少實驗種類

需要隨時organize data and perform statistical analysis? • 有點耗時，但可幫忙還須多少次實驗以達到統計上有意義之差異。 • Data的意義常不是第一眼便看出。 • 計算你平均耗多少時間於(1) design experiment (2) perform experiment (3) analyze the data

數據不符advisor想法被要求反覆重複 • Understand the rationale of advisor’s idea. We do not reject a hypothesis with a simple experiment. On the other hand, the data from different lab person must be coherent. • Adjust the experimental details to get the data to fit in the hypothesis with integrity. • People usually try the “best” effort on their own hypothesis. The advisor also need to try convince the student.

堅持原有之想法或改換新方向？ • We may not get the “idea” data to demonstrate our hypothesis or we may find another “novel and ”interesting” hypothesis as your new research direction. • Changing idea is not the default pathway. • 成功的人會一直往前走，雖然不免犯錯，但絕不輕言放棄—Conrad Hilton

老闆是要培養學生創造力還是太忙沒空理學生？老闆是要培養學生創造力還是太忙沒空理學生？ • Do not frequently ask “what do I do next? • Be sure that you must accept the responsibility for managing the relationship between you and your advisor. Arouse the interest of your advisor by novel and solid result. • Help your supervisor to give you better feedback on your work. Always ask supplementary questions to ensure that you understand fully what is being required of you. • You need to educate your supervisor continually. Especially, on the research topic, in which you are fast becoming the expert.

學生須要自己寫 paper to final format? • Rationale is not carved on stone. • Reorganization of data to fit in the question you address in the paper. • Do not arrange the data chronologically in the thesis or paper. • The aims of the paper may not be the original aim of your research.

如何寫一個簡單計劃書 • Virtual research plan (simulation of a real research project) • 任卓潁said: find the title of an article, and try to plan the experiment to get the result before reading the abstract and full-text.

聽seminar有益博士班學生研究？ • Research seminar can provide (1) what is the current research trend (2) What type of experimental proof is required (3) What type of material is available (4) new concept for your own experiment. • Listen to the seminar you are interested in, and attend the seminar outside your field sometimes.

須取得老闆注意或重視？ • In the lab, there always seems to be some one who has the P.I.’s attention and admiration. Look carefully, Is it deserved? Maybe you can learn something. And if it is not deserved, mind your own business and learn not let it bother you. • Today’s up and comings may be tomorrow’s down and outs.

實驗室人員相處不佳 • The inconsiderate lab researcher is always a problem to many laboratories. • You may need to examine your personal attitudes when more than two persons out of ten dislike you.

如何合法由台北去台東而不花一毛錢？ • 找搬家公司正由台北往台東市，先幫他們搬家具上車，再請求順便搭便車到台東。 • 創造他人價值：make yourself be useful to others. • 參加○明德之環島倒○車隊活動 • 了解周圍發生之事：Do not mind your own experiment only.

Four Advices • Aim and Enthusiasm • SWOT (Strength, Weakness, Opportunity, and threat) • Be well-prepared through virtual training • Work hard and Leave the rest to God

作為一個學者已經犧牲很多事情不要怕 To be yourself陳明哲，維吉尼亞大學達頓商學院講座教授

Muhammad Yunus • Every single individual on earth has both the potential and the right to live a decent life. Across cultures and civilizations, Yunus and Grameen Bank have shown that even the poorest of the poor can work to bring about their own development. • The Nobel Peace Prize 2006

References 1. “Advice to a young scientist” by Peter Medawar, 1981. 2. “At the bench, a laboratory navigator” by Kathy Barker, 1998. Cold Spring Harbor Laboratory. 3. “Who wants to be a scientist” by Nancy Rothwell.中譯本：科學家的第一堂課，陳佳伶譯，早安財經文化有限公司，2004。

一些生醫研究的迷思 A few paradox in biomedical research