DNA extraction quantitation DNA 50 ngL DNA PCR

今日上課流程 • • 課程回饋 (實驗二 - DNA extraction & quantitation) 開始上課 DNA 濃度稀釋 (50 ng/µL) DNA 濃度檢測 PCR 試劑配置 (Target gene: Lectin & 35 s promoter) PCR 電泳跑膠

除草劑 - Round up

Transgenic Soybean gene Mechanism of Roundup (Glyphosate)

Transgenic Soybean gene Mechanism of Roundup (Glyphosate)

Transgenic Soybean gene

polymerase chain reaction (PCR)

PCR principle

PCR principle

PCR principle

PCR principle-Primers

DNA extraction

Gel electrophoresis of soybean PCR lectin gene

Gel electrophoresis of soybean PCR 35 S promoter gene

Gel electrophoresis of soybean PCR EPSP gene

Primers

DNA 濃度稀釋 (50 ng/µL) 組別 初始濃度 (ng/µl) 1 3028. 56 60. 57 595. 7 2 515. 26 10. 30 93 3 1663. 14 33. 26 322. 6 4 2794. 46 55. 88 548. 8 5 2056. 49 41. 13 401. 3 6 406. 52 8. 13 71. 3 7 530. 55 8 3958. 96 79. 17 781. 7 9 1122. 89 22. 45 214. 5 10 1644. 19 32. 88 318. 8 11 1430. 87 28. 60 276 12 1608. 23 32. 16 311. 6 13 153. 72 3. 07 20. 7 最終濃度 50 ng/µL 稀釋倍數 10. 61 DNA 量 10µL 補水 (µl) 96. 1

DNA 濃度檢測 • Nanodrop 使用方 法 Main manu Water or 2µL DNA Sample ======= 1. 2. 3. 4. ds. DNA ss. DNA RNA Protein ======= == Function ===== Data ==== ---- ========== 1. Select sample 2. Start Measurement 3. Show & Print Results 4. Time Setting 5. About Maestrogen ========== Select sample Blank Sample(ds. DNA) Data Save & Exit * * Blank Sample_01 Sample_02 Sample_03 =========== == Function ===== Data ==== 註: 每個樣品檢測完用拭鏡紙擦拭 檢測結束用純水洗淨並擦拭乾淨 ---- Select All Select Print Data Menu Return * Blank * Sample_01 * Sample_02 ===========

PCR 試劑配置 Target gene: 35 S Master mix Target gene: Lectin 10µL Master mix (Taq, buffer, d. NTP, dye) H 2 O Primer Soybean 35 S F Soybean 35 S R (Taq, buffer, d. NTP, dye) 1µL H 2 O Primer Lectin F Lectin R 4µL 19µL 1µL DNA G 1 …………. G 13 10µL 1µL 4µL 19µL G 1 …………. G 13 1µL DNA 註：在蓋子上寫組別和 Target gene

PCR conditions • Denature 94℃ 2 min 94℃ 30 sec 55℃ 1 min 35 Cycle 72℃ 1 min • 72℃ 2 min

助教預做結果 14 K M A Lectin B C D 35 S promoter A B C D

各組黃豆種臍顏色 G 1 G 2 G 6 G 7 G 11 G 3 G 4 G 5 G 8 G 9 G 10 G 12 G 13