Teacher蔡: (1) Fig 1C western 太髒, not qualified for publication (要改進!). (2) EAD clone 88 is a good one for IFA (3) Akata strain 比 B95-8 transforms B cell 能力弱很多. (3) Dox-treated ERKV中 knock-downKR (siRNA),用以證實ER是透過KR做事... (4) ACIF是做好EBNA1 IFA之唯一良藥. (5)淑君的Blood paper中述及primary infection可有Z, R, 一直到VCA. (6)葉醫師的differentiation系統為Sphenoidal sinus, 並未有un-differentiated cells.
Teacher陳M.-R.: (1) MA 220 is a good antibody for Dox-EREV8 late gene expression (又是Fig 1C!) (2) Mechanism of ER-mediated activation of KR promoter.
Teacher陳J.-Y.: 要做與疾病,癌症有關的為佳!
SFL: (1) 一篇會令人開悟的文章, 這裡.(2) 293的Vitronectin receptor是怪胎 (ITGAVB1). (3)NA中ER的量與EREV8中ER的量是可以比較估算的.
Jan 28, 2010
Jan 22, 2010
B292 相關
1/21/2010 (Thu) 第一次討論會議
Speaker 1-北醫劉景平: Structures of 292, J-30, SAHA etc. One OCH3 group in 292 is enough, two OCH3 increases IC50 from 50 nM to 1400 nM; replace with OH increases IC50 from 50 nM to 500 nM. SAHA is a planer structure, quite stable. SAHA is a pan-HDACi, with uM range. B291 is also a pan-HDACi, with 100 nM range. Wishing 292 is more specific.
Speaker 2-程韻靜: Effects of 292 (1/3) inhibits tubulin polymerization in KB cell; (2/3) inhibits tube formation in SH-SY5Y; (3/3) Animal models (CAM, xenograft in H460 58.7% inhibition, orthotopic PANC-1 with Luc expression); Calcium influx was increased in Taxol or J-30 (not as good as Taxol) treated neuronal cells, but neither SAHA nor 292 had effects in calcium influx; While most cells exhibit 50~100 nM IC50, HUVEC is 42 uM and WI-38 is 12 uM (so interesting!!!)
Speaker 3-夏興國: (1) B292 and EMT seemed not have correlation in breast and lung cancer cells (2) B292 and Adhesion: CL1-0 扁平+292 (uM) =扁平. CL1-5 細長+292 (nM)=morphological changes and tubulin disperse.... (但butyrate, TSA, SAHA都能造成相同現象(FAK and paxillin increases) 耶...
Speaker 4-吳仲峻: TWO1 (72 nM), NA (35 nM with EAD 5~10%), HONE-1 (66 nM), HA (42 nM).
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11/1/09 SFL wrote:
與PrCA/AR有關:Taken together, our studies suggest that HDAC6 regulates AR hypersensitivity and nuclear localization, mainly via modulating HSP90 acetylation.
與Tau有關:To determine the relationship of tau protein and increased tubulin acetylation, we studied the effect of tau on the acetylation-deacetylation of tubulin. Our results indicate that tau binds to the tubulin-deacetylase, histone deacetylase 6 (HDAC6), decreasing its activity with a consequent increase in tubulin acetylation.
有趣, 有趣..
有空要瞭解一下HSP90,就這麼查吧. "hsp90 review AND cell cycle [jour]"
SAHA 相關 Chiang's recent paper [Here].
Speaker 1-北醫劉景平: Structures of 292, J-30, SAHA etc. One OCH3 group in 292 is enough, two OCH3 increases IC50 from 50 nM to 1400 nM; replace with OH increases IC50 from 50 nM to 500 nM. SAHA is a planer structure, quite stable. SAHA is a pan-HDACi, with uM range. B291 is also a pan-HDACi, with 100 nM range. Wishing 292 is more specific.
Speaker 2-程韻靜: Effects of 292 (1/3) inhibits tubulin polymerization in KB cell; (2/3) inhibits tube formation in SH-SY5Y; (3/3) Animal models (CAM, xenograft in H460 58.7% inhibition, orthotopic PANC-1 with Luc expression); Calcium influx was increased in Taxol or J-30 (not as good as Taxol) treated neuronal cells, but neither SAHA nor 292 had effects in calcium influx; While most cells exhibit 50~100 nM IC50, HUVEC is 42 uM and WI-38 is 12 uM (so interesting!!!)
Speaker 3-夏興國: (1) B292 and EMT seemed not have correlation in breast and lung cancer cells (2) B292 and Adhesion: CL1-0 扁平+292 (uM) =扁平. CL1-5 細長+292 (nM)=morphological changes and tubulin disperse.... (但butyrate, TSA, SAHA都能造成相同現象(FAK and paxillin increases) 耶...
Speaker 4-吳仲峻: TWO1 (72 nM), NA (35 nM with EAD 5~10%), HONE-1 (66 nM), HA (42 nM).
------------------------------------------------------------------------------
11/1/09 SFL wrote:
與PrCA/AR有關:Taken together, our studies suggest that HDAC6 regulates AR hypersensitivity and nuclear localization, mainly via modulating HSP90 acetylation.
與Tau有關:To determine the relationship of tau protein and increased tubulin acetylation, we studied the effect of tau on the acetylation-deacetylation of tubulin. Our results indicate that tau binds to the tubulin-deacetylase, histone deacetylase 6 (HDAC6), decreasing its activity with a consequent increase in tubulin acetylation.
有趣, 有趣..
有空要瞭解一下HSP90,就這麼查吧. "hsp90 review AND cell cycle [jour]"
SAHA 相關 Chiang's recent paper [Here].
Jan 16, 2010
Jan 7, 2010
Dec 21, 2009
Dec 18, 2009
新的挑戰—for YCK (更新)
真是不好意思,我又改變順序了
1. 回溶HeLaTet (#3、#4)及HONE1Tet
2. 測試Bsd在TW03、CGHNK2及Zeo對HeLaTet、HONE1Tet的耐受性
3. 293FT用來包pLentiTR6, pLentiFlag-gZ, -cZ及GFP (每種包3個well, 就是半盤6-well plate之意).
同時,可否問一下進階業務,"LONZA"有款叫KGM的medium,專門用來養keratinocyte,是否有進一部資料可供參考! 謝謝.
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on 12/2/2009 Crazy SFL wrote:
哈哈哈, 新年新氣象,這是給YCK和YLC的小禮物。先把手邊的東西忙完後,請著手下面有趣的實驗:
1. 回溶HeLaTet (#3、#4)及HONE1Tet
2. 測試Bsd在TW03、CGHNK2及Zeo對HeLaTet、HONE1Tet的耐受性
3. 293FT用來包pLentiTR6, pLentiFlag-gZ, -cZ及GFP (每種包3個well, 就是半盤6-well plate之意).
同時,可否問一下進階業務,"LONZA"有款叫KGM的medium,專門用來養keratinocyte,是否有進一部資料可供參考! 謝謝.
----------------------------------------------------------
on 12/2/2009 Crazy SFL wrote:
哈哈哈, 新年新氣象,這是給YCK和YLC的小禮物。先把手邊的東西忙完後,請著手下面有趣的實驗:
- Try to establish TW03-TetR, CGHNK2-TetR, CGHNC8-TetR and TW2.6-TetR。
- Try to establish Luc, ER and gZ (B95-8 strain) in HONE1Tet, NP460Tet, TW03Tet, CGHNK2Tet, CGHNC8Tet and TW2.6Tet。這裡有個重點就是請再重讀一下原instruction manual, 把induction量弄到和293TetER相可匹配的程度。我們需要步步為營。
Dec 1, 2009
量化西方墨漬法
以後實驗室quantitate protein band請依以下步驟進行:
- Plasmid 最好是一起抽的一批, 因實驗室LB/BHI已過使用期限,希望大家每次抽質體時盡量使用新鮮media及LB plate (1-cm厚度最佳),超過一個月以上的media及plate請自行定時清理(請不要當福壽螺到處蔓延留種,謝謝)。
- 若質體中有SV40-ori, 請以293T為host (DLR/expression etc),其餘它種質體則用293。
- Transfection (24~48 h)每次請做p-100/1 ml-lysate的份量(6x p-35/1.2-ml), protein lysate量一次到位,不要老是重複在養細胞/TF效率好像有問題/plasmid不夠...(姑且稱之為小鼻子小眼睛的實驗)
- 檢測各質體間表現量:(i) 請以wild-type為標準 (所以wild-type可能要多transfect一份),293T為host的取5-ug及10-ug, 293為host的取25-ug及50-ug protein extracts run數片小膠,確定所使用的西方墨漬條件可分出一倍和兩倍間的差異。若是過強,那就請在砍primary/ secondary antibody濃度、incubation時間、ECL強度上三件事情上做調整。(ii)依前步驟所訂出的條件run一片小膠檢查各檢體間是否差異很大,如有二倍以上差異而各步驟又沒有太大疑慮,那表示可能有protein stability/proteosome involved,值得設計其他實驗來分析之.兩倍以下差異者微調input量可繼續往下的實驗。(Plasmid, 細胞用畢的請定時清理,不要當福壽螺到處蔓延留種,謝謝)。
- 附註定量software有三款,請在Lab Protocol中找到小抄。
Novartis 石明石博士 Michael Shi 演講
- TKI 258 was a FGFR1 inhibitor, but the use of TKI 258 causes FGF23/bFGF increased.
- Melanoma trial found FGF mediated escape form anti-angiogenesis (sorxxxtinb/...) Early phase: VEGFR2 blockadge.
- Phase 3 trials: (a) FGFR3 expression/t94,14) trans-location in melanoma (b) FGFR3 mutations in bladder cancer (c) FGFR1 expression in breast cancer [這裡若有蒲教授的data就好了!!]
- AUY 922 is an inhibitor of HSP90 for GI tract tumor.
- BGT226 is is on clinical trial for NPC (與中文大學馬教授合作,明年AACR發表) 另一位Mike補充陳所長有拿藥用在oral cavity (張光裕醫師 was in charge, he found this drug induced autophagic death in a panel of oral cancer cells)
- BKM120 is a PI3K -specific inhibitor; BGT226 and BE2 232 are dual kinase inhibitors (mTOR and PI3K) 這部份還是沒有搞清楚.
- ASA 404 in NSCLC.
- LED 225 is for Smoothened/Hedgehog pathway.
- MGM 黃秀芬醫師介紹 Tissue bank/LCM slides for Chlorangiocarcinoma...
Nov 21, 2009
知識分享 - immunopreciptitation
LHC大型強子對撞器→修好了
Nov 19, 2009
Nov 18, 2009
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