(3.236.231.61) 您好!臺灣時間:2021/05/11 21:03
字體大小: 字級放大   字級縮小   預設字形  
回查詢結果

詳目顯示:::

我願授權國圖
: 
twitterline
研究生:李至芬
研究生(外文):Chih-Fen Li
論文名稱:昆蟲細胞株檢定蘇力菌內毒素毒效之研究
論文名稱(外文):Determination of Bacillus thuringiensis δ-endotoxin with insect cell lines
指導教授:侯豐男侯豐男引用關係
指導教授(外文):Roger F. Hou
學位類別:碩士
校院名稱:國立中興大學
系所名稱:昆蟲學系
學門:生命科學學門
學類:生物學類
論文種類:學術論文
論文出版年:1995
畢業學年度:83
語文別:中文
論文頁數:55
中文關鍵詞:蘇力菌內毒素昆蟲細胞株快速檢定系統
外文關鍵詞:Bacillus thuringiensisendotoxininsect cell linesrapid detective system
相關次數:
  • 被引用被引用:0
  • 點閱點閱:137
  • 評分評分:系統版面圖檔系統版面圖檔系統版面圖檔系統版面圖檔系統版面圖檔
  • 下載下載:0
  • 收藏至我的研究室書目清單書目收藏:0
本試驗以3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium
broide(MTT)細胞檢定法測定蘇力菌(Bacillus thuringiensis; Bt)內毒
素毒害細胞之程度。主要是利用MTT與細胞內粒線體中去氫酵素作用後之
呈色反應, 而算出細胞數目, 進而推算出細胞毒性(cytotoxicity)百分率
。不同蘇力菌品系之活化內毒素對不同細胞株的毒效測定, 以PX- 1187細
胞之毒性較優於其他細胞株, 而SF-21AE、SF9及TN-368細胞對Bt內毒素之
感受性較低。測定不同Bt品系所純化之內毒素對小菜蛾四齡幼蟲的半數致
死濃度(LC50), 試驗結果顯示, B. t. subsp. kurstaki(Btk) HD1及
HD263之效果最佳, 其次為Btk HD73, 再者為Bt0158,而結果與in vitro之
檢定結果相符。進一步以Btk HD263之活化內毒素篩選PX-1187細胞, 篩選
初期隨著篩選次數的增加, 細胞對Btk HD263之感受性逐漸降低。篩選至
第九代(PXR-HD263-9),細胞對Btk HD263之感受性降至15.14%, 且至第20
代(PXR-HD263-20), 細胞死亡率並無明顯的變化。以cBtk HD73之活化內
毒素篩選PX-1187細胞, 也得到類似的情形; 而以Btk HD263活化內毒素處
理小菜蛾B單株及F單株細胞, 則無細胞死亡率逐漸下降的現象。當PXR-
HD263細胞不再以內毒素處理, 細胞對Btk HD263之感受性逐漸回升。以聚
合酵素連鎖反應(polymerase chain reaction, PCR)鑑定各蘇力菌品系所
含的cry I基因。結果Btk HD1、HD73及HD263均含有cryIA(c)基因。不同
的蘇力菌品系可能含有相同或相似的晶體毒蛋白(insecticidal crystal
proteins, ICPs), 可能為細胞對蘇力菌品系造成交互抗性的主要原因。

A rapid detective system has been developed to assese the
activity of toxins produced by Bacillus thuringiensis (Bt) on
insect cell lines. The assay was performed in a 96-well plate
using MTT as the substrate. The compound is reduced by
mitochondrial dehydrogenases of live cells to exhibit a purple
color. The amount of dye converted is proportional to the
number of living cells and then δ-endotoxin cytotoxicity can
be calculated. Based on determination of cytoxicity to viable
insect cells, PX-1187 and BmN cell lines were found sensitive
to endotoxin of B. t. subsp. kurstaki(Btk) HD1、HD73、HD263 and
B. t. subsp. israelensis(Bti) IPS82. Toxins of Bt0158 and NTU9
were significantly toxic to PX-1187 cell line. But SF-21AE、SF9
and TN-368 cell lines were insensitive to these pure toxins of
Bt strains. The LC50 values of Bt endotoxin against 4th-instar
larvae of P. xylostella was calculated. For all six Bt strains,
HD1 and HD263 were the most toxic, followed by HD73 and Bt0158.
These results were similar to those in vitro tests. On the
other hand, we selected for the PX-1187 resistant cells by
subculturing in the presence of HD263 toxic proteins. The
resistant cell could be readily selscted from a mixed cell
population by cultivation in the presence of δ-endotoxin.
Cytotoxicity of HD263 against PXR-HD263-9 cell achieved
15.14%. PX-1187 cells can be similarly selected for resistant
to HD73 toxic proteins.

QRCODE
 
 
 
 
 
                                                                                                                                                                                                                                                                                                                                                                                                               
第一頁 上一頁 下一頁 最後一頁 top
系統版面圖檔 系統版面圖檔