附表六 長庚大學學生出席國際會議報告書 填寫日期: 98 年 4 月 30 日 報告人姓名 陳泰龍 系所及年級 生物醫學所生技組博一 學號 D9701412 會議期間 2009/4/16~4/17 核定補助日期 98 年 3 月 16 日 會議地點 美國馬里蘭州 巴爾地摩 會議名稱 (中文) 2009 第 41 屆橡樹山陵年會 (英文) 41st Annual Oak Ridge Conference - 2009 發表論文題目 (請附論文全文) (中文) 利用肽鏈核酸螢光探針偵測有抗藥性(抗濾過性病毒劑) 且微量存在的 A 型流感病毒 (英文) Use of peptide nucleic acid probe for homogenous detection of low abundant, Amantadine-resistant influenza A virus 內容摘要 Detection of drug-resistant mutant of pathogens is important for a clinician to choose an appropriate therapeutic strategy. However, to detect such mutants is usually difficult as they might only exist in a low abundance in the population before treatment. In this study, we have developed a novel method combining an anchor probe and a fluorophore-labeled peptide nucleic acid (PNA) probe that covered the mutation hotspot of matrix protein 2 gene of influenza A virus for homogenous detection of Amantadine-resistant virus. PNA is a DNA analog in which the phosphodiester backbone is replaced with an N-(2-aminoethyl)glycine chain. A wild-type specific PNA oligomer can selectively inhibit the amplification of wild-type template but allow the amplification of mutant templates in polymerase chain reaction. The fluorophore-labeled PNA was also designed as a sensor probe to reveal the melting curve profile of the PCR products. By the melting curve analysis after PCR amplification, the mutant could be identified through its signature melting peak. On a LightCycler machine, this assay can be finished in a single tube in less than 30 minutes. The detection limit of this assay is less than 10 copies of mutant viral genome in the existence of 1000 fold excess of wild type genome. Using this method, we
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附表六
長庚大學學生出席國際會議報告書
填寫日期: 98 年 4 月 30 日
報告人姓名 陳泰龍系所及年級 生物醫學所生技組博一
學號 D9701412
會議期間2009/4/16~4/17
核定補助日期 98 年 3 月 16 日
會議地點美國馬里蘭州
巴爾地摩
會議名稱 (中文) 2009 第 41 屆橡樹山陵年會
(英文) 41st Annual Oak Ridge Conference - 2009
發表論文題目
(請附論文全文)
(中文) 利用肽鏈核酸螢光探針偵測有抗藥性(抗濾過性病毒劑)
且微量存在的 A型流感病毒
(英文) Use of peptide nucleic acid probe for homogenous detection of
low abundant, Amantadine-resistant influenza A virus
內容摘要
Detection of drug-resistant mutant of pathogens is important for a
clinician to choose an appropriate therapeutic strategy. However, to
detect such mutants is usually difficult as they might only exist in a
low abundance in the population before treatment. In this study, we
have developed a novel method combining an anchor probe and a
fluorophore-labeled peptide nucleic acid (PNA) probe that covered
the mutation hotspot of matrix protein 2 gene of influenza A virus
for homogenous detection of Amantadine-resistant virus. PNA is a
DNA analog in which the phosphodiester backbone is replaced with
an N-(2-aminoethyl)glycine chain. A wild-type specific PNA
oligomer can selectively inhibit the amplification of wild-type
template but allow the amplification of mutant templates in
polymerase chain reaction. The fluorophore-labeled PNA was also
designed as a sensor probe to reveal the melting curve profile of the
PCR products. By the melting curve analysis after PCR
amplification, the mutant could be identified through its signature
melting peak. On a LightCycler machine, this assay can be finished
in a single tube in less than 30 minutes. The detection limit of this
assay is less than 10 copies of mutant viral genome in the existence
of 1000 fold excess of wild type genome. Using this method, we
have successfully identified 10 mutants from 21 amplifiable, throat
swap samples collected from patients before treatment, which is in
agreement with the results using traditional methods involving viral
culture, PCR, and sequencing. However, we did not find mutant
quasi species in the wild type population, indicating that the
Amantadine-resistant mutant may not co-exist with the wild-type
virus in most patients, or that the mutant exists in an even lower
ratio than the detection limit. In summary, we have developed a
rapid and sensitive method using PNA probe to enrich and detect
low abundant viral mutants. This method may also be useful for the
detection of rare drug-resistant mutants in other pathogens and
cancer cells.
一、報告內容請另頁撰寫,以 A4、標楷 12、橫寫、直式列印,內容包括:
(一)、參加會議經過
(二)、與會心得(以獲取專業新知為主,其他新知,例如人生經歷等為輔)
(三)、建 議
(四)、攜回資料名稱與內容
(五)、其 他
二、系所安排口頭心得報告時間:98 年 6 月 16 日
(未安排原因:_________________________)
研發長
所長
指導教授
報告人
學生出席國際會議報告書及網路刊載由研發處備存辦理。
(一)、參加會議經過
本次會議在美國馬里蘭州的巴爾地摩市舉行,巴爾地摩位於華盛頓 DC 的東
北方車程約一小時處,會議名稱為 Oak Ridge Conference,由美國臨床化學學
會(American Association for Clinical Chemistry, AACC)主辦,會議時間是