熱逆境對小球藻染色體與細胞膜的影響

The effects of heat stress on chromosomes and cell membranes of chlorella

張晉嘉

指導教授 ： 徐邦達

國立清華大學/生命科學院/生物資訊與結構生物研究所/碩士(2006年)

https://doi.org/10.6843/NTHU.2006.00253

摘要

小球藻經過46.5℃加熱一小時之後分別在照光與避光的兩種環境下培養，其型態和生理都出現極大的差異。照光的細胞會白化，但避光的細胞則一直維持綠色。利用核酸染劑(DAPI和Acridine orange)在共軛焦顯微鏡下可以觀察到照光細胞的DNA濃縮得很嚴重，而避光細胞的DNA濃縮的程度並不像照光劇烈。在葉綠素方面，照光細胞的葉綠素在十小時內幾乎分解消失；避光細胞的葉綠素一直都存在著，但無光合作用的活性。本研究為了更深入了解熱逆境後照光與避光的差異，再使用SYBR Green 1和SYTOX Green兩種染劑來分析之間的不同；並以Evans blue和Trypan blue比較照光與避光細胞膜破損的情形，發現光照的小球藻細胞膜破損的數量遠遠高於避光的小球藻，且細胞膜破損的細胞高達60%以上。由以上實驗，可以更確定光照所帶給小球藻的傷害是存在的，且不論是照光或避光，DNA均不會分解消失。而這其中的生理機制，值得更深入研究。

關鍵字

小球藻 ； 核酸染劑 ； 細胞膜 ； 共軛焦顯微鏡 ； 流式細胞儀

並列摘要

After heating chlorella pyrenoidosa at 46.5℃ for an hour, and culturing them under light or in the dark regime, we found that there are quite difference between chlorella’ pattern and physiology on result. The cell under light regime is etiolating and the cell under dark regime is kept green.
We used the DAPI and Acridine orange to observed the cell by Confocal Microscopy, we can see the DNA of the cell in light regime were condensed seriously but the cell under dark regime is not seriously. About the pattern of the chlorophyll; the chlorophyll of the cell in the light regime almost disappear in 10 hours, on the other hand the chlorophyll of the cell in the dark regime existence to the end but no photosynthesis activities.
Other two fluorescence stains, SYBR Green 1 and SYTOX Green are also used to analyze the difference under between light regime and dark regime. Evans blued and Trypan blue to compare how the rupture situations of cell membrane were. Here we found out the number of rupture chlorella in light regime is much higher than dark regime, also the percentage of rupture cell more than 60%.
According to result we obtained, we know the light could cause injured of the chlorella and no matter in light regime or dark regime, the DNA won’t dissolution and disappear.
We think that the physiology mechanism worth studying deeply in chlorella of the situation in the future.

參考文獻

Bernardi P.,Scorrano L.,Raffaele C.(1999)Mitochondria and cell death – mechanistic aspects and methodological issues. Eur.J.Biochem. 264:687-701

Bewley J.D.(1979)Physiological aspects of desiccation tolerance. Ann.Rew.Plant Physiol. 30:195-238

Briggs C. and Jones M.(2005)SYBR Green 1-induced fluorescence in cultured immune cells:A comparison with Acridine Orange. Acta histochemica 107:301-312

Buckner B.,Johal G.S.(2000)Cell death in maize. Physiol.Plantarum 108:231-239

Crowe J.H.,Crowe L.M.,Carpenter J.F. and Wistrom C.A.(1987)Stabilization of dry phospholipid bilayers and proteins by sugars. Biochem.J. 242:1-10