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標題: | 溫度對吳郭魚細胞紫外線敏感度和基因表現的影響 I. Temperature Effects on DNA Replication and UV Repair in Tilapia Ovary Cells. II. Thermal Adaptation and Heat Shock Response of Tilapia Ovary Cells |
作者: | Jerng-Don Chen 陳振當 |
關鍵字: | fish cells,temperature,UV sensitivity,DNA replication,excision repair, |
出版年 : | 1987 |
學位: | 碩士 |
摘要: | 本實驗的目的是探討溫度對吳郭魚卵巢細胞(TO-2 cells)紫外線(UV)敏感度和熱休克蛋白質(heat shock proteins, hsp)基因表現的影響。吳郭魚卵巢細胞株建立於1983年,此細胞可以在廣溫度的環境下生長(15℃?34℃),且其對UV的忍受性比哺乳類細胞還強,這一特性與其他魚類細胞不同,因為大部份的魚類細胞對UV都是相當敏感的。在此我們探討TO-2細胞在不同溫度下對UV的敏感度、DNA複製和UV修復能力的改變;並研究吳郭魚細胞獲得抗熱性的能力,以及溫度改變對其熱休克蛋白質基因表現的作用情形。 UV會抑制DNA的合成速率,此一抑制作用在37, 31和25℃下3小時內能夠恢復到正常,然而在18℃下則否。當細胞生長在低溫(18℃)的時候DNA的合成速率減慢,而在高溫狀態下(37℃), DNA的合成速率一開始先下降至60%,且主要的抑制是在DNA合成的起始步驟(initiation), 8小時後則回到比正常還高的速度(180%)。DNA合成的起始步驟在4J/m2的UV照射下會受到抑制,到9J/ m2時,大分子量的DNA合成中間產物也開始受到抑制,而小分子量的DNA中間物則會產生堆積,此一堆積起的小分子DNA可以被延長成高分子,顯示其具有複製後修補(postreplication repair)的能力。在37℃下,則需要較高劑量的UV照射方能影響其DNA的合成。 TO-2細胞對UV的切除修復能力(excision repair)並不高,但卻是目前所知道魚類細胞中最好的。在不同的溫度下,我們測量UV照射所引的雙嘧啶環(pyrimidine dimers)的切割速率(incision rate),並計算切割酵素作用所需的活化能(activation energy)。 我們並探討TO-2細胞在不同溫度下的增殖、大小、以及細胞週期的分佈情形,並研究其抗熱性(thermotolerance)和熱休克蛋白質基因表現的相關性。低溫處理(18℃或4℃)對TO-2細胞並無明顯的致死作用,且細胞大小及細胞週期亦無明顯變化;然而在高溫(37℃)處理3天後,只有1%的細胞存活,且細胞的體積逐漸增大,並有明顯的G2時期延長現象。 當細胞先以一溫和的熱處理後,其對下一次致死熱處理便具有抵抗性。TO-2細胞可由37℃的慢性熱處理(chronic heating)或40℃、15分鐘的短暫熱處理(acute heat shock)而獲得對下一次43℃熱處理的抵抗性,且此一抗熱性的獲得伴隨著熱休克蛋白質基因的過量表現(over expression)。 TO-2 is a fish cell line derived from Tilapia ovary. It grows in a wide range of temperature (15°-34℃). The cells are more UV resistant than mammalian cells; they are distinguished from other cultured fish cells, most of which are UV sensitive. In this report we present the effects of temperature on UV sensitivity, DNA replication, and UV repair on TO-2 cells. UV suppressed but could not block DNA synthesis completely. The inhibition was overcome in 3 hours at 37°, 31°, and 25℃, but not at 18℃. Initiation of nascent DNA synthesis was blocked at 4 J/m2 in TO-2 cells compared with <1 J/m2 in mammalian cells. After 9 J/m2 UV irradiation, low molecular weight DNA replication intermediates started to accumulate, and they could be chased into high molecular weight DNA with little delay. Cells grown at low temperature (18℃) showed reduced rate of DNA synthesis. Upon shifting to sublethal high temperature (37℃), the rate first decreased to 60 percents by suppressing initiation and after 8 hours at 37°C the rate of DNA synthesis overshot and reached 180 percents of the control. TO-2 cells showed low level of UV induced excision repair; but this was prominent as compared with other fish cells. The incision rate of pyrimidine dimers at various temperatures have been measured, and the activation energy of incision estimated. Keywords: fish cells, temperature, UV sensitivity, DNA replication, excision repair |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/75597 |
全文授權: | 未授權 |
顯示於系所單位: | 動物學研究所 |
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