복합 식물 총 RNA (...을 더한) 소량 추출 키트

1. 시약 키트의 구성 요소

2. 저장

This reagent kit can be stored at room temperature (15-25℃) 건조한 환경에서 안정적이며 12 개월.

3. 시약 키트 사용 지침

3.1 이 키트는 분자 생물학 연구 목적으로 제작되었으며 질병 진단이나 치료에 사용해서는 안 됩니다..

3.2 키트의 일부 구성품에는 자극제가 포함되어 있습니다.; 필요한 예방 조치를 취하는 것이 좋습니다 (보호복과 고글을 착용하는 등).

3.3 이 키트를 사용하려면 고속 원심분리기 등 추가 장비가 필요합니다., 욕조 (금속 욕조), 소용돌이 믹서, 무수에탄올, 액체 질소, 클로로포름, 멸균 탈이온수, 및 EP 튜브.

4. 시약 키트 소개

This RNA purification reagent kit provides a fast and effective purification of plant total RNA containing polysaccharides, lipids, polyphenols, and other components. It is suitable for most complex plant tissues. In general, lipid-rich plant tissues contain a high content of lipid compounds, which significantly affect RNA extraction efficiency. This kit utilizes exclusive inhibitor removal columns to effectively eliminate lipids from plant samples, as well as to remove DNA contamination. If the experiment is sensitive to DNA, it is recommended to use intron-spanning primers for downstream experiments.

The RNA rapid purification reagent kit can extract plant total RNA (핵 RNA 및 세포질 RNA 포함) 이내에 1 시간. 추출된 RNA를 RT에 바로 활용 가능-PCR, 노던 블로팅, 등. The entire purification process does not require toxic reagents such as chloroform, making the RNA purification reagent kit suitable for various other samples.

5. 실험 원리 및 절차

6. 추출과정

실험 시작 전 주의사항:

ㅏ. 씻다 완충기 1: 사용하기 전에, add the specified amount of absolute ethanol as indicated on the reagent bottle label. Check the label to confirm the addition of absolute ethanol.

비. 용출 완충액은 0.1x TE 솔루션 최소한의 EDTA로. EDTA가 후속 실험에 영향을 미칠 수 있는 경우, it is recommended to replace the elution buffer with sterile deionized water.

씨. RNA Extraction Buffer I contains a small amount of phenol, which may precipitate. 사용하기 전에, mix thoroughly by warming in a water bath; after use, store away from light.

1. 샘플 처리:

ㅏ. 재료 수집 및 보관:

If freshly collected materials cannot be immediately used, place them in liquid nitrogen and store them at -80℃.

비. Whenever possible, collect fresh materials as they contain fewer polysaccharides and polyphenols.

2. Grind approximately 100 mg of fresh samples or not more than 20 mg of dry material with liquid nitrogen.

3. 추가하다 600μl of RNA Extraction Buffer I, ensuring there are no tissue clumps in the ground sample. Tissue clumps are difficult to lyse and can reduce RNA yield.

4. Vortex for 30 에스.

5. Transfer the lysate to aninhibitor removal purification column, 원심분리기 12,000 rpm 5 분.

(메모: Lipid-rich plant materials may contain many lipid compounds at this step, which can affect RNA extraction. Remove these substances during this step. If buffer residue remains in the inhibitor removal purification column during centrifugation, extend centrifugation time appropriately.)

6. Transfer the obtained supernatant to a DNA removal column, 원심분리기 12,000 rpm for 30s, and collect the filtrate (메모: RNA is present in the filtrate).
7. 추가하다 250무수 에탄올 μl, mix by pipetting. If there is a small amount of precipitation, it does not affect subsequent experiments. Transfer the liquid to an RNA purification column, 원심분리기 12,000 rpm for 30s, discard the flow-through.
8. 추가하다 700μl of inhibitor removal buffer, 원심분리기 12,000 rpm for 30s, discard the flow-through.
9. 추가하다 700μl의 washbuffer 1 RNA 정제 컬럼으로, 원심분리기 12,000 rpm for 30s, discard the flow-through.
10. 추가하다 500μl의 wash buffer 1 RNA 정제 컬럼으로, 원심분리기 12,000 rpm 3 분, discard the flow-through.
11. Place the RNA purification column into a new 1.5ml centrifuge tube, air-dry the membrane at room temperature for 2 분.

(메모: Confirm that absolute ethanol has been added to wash buffer 1. 에탄올의 존재는 후속 실험에 심각한 영향을 미칩니다., so drying the membrane is crucial. 원심분리 후, 용출 전에 에탄올이 존재하지 않는지 확인하십시오., 그런 다음 폐기물 및 수집 튜브를 폐기하십시오.. After washing with wash buffer 1, the membrane on the RNA purification column should have only a slight color. 원심분리 후, carefully remove the RNA purification column, ensuring it does not touch the collection tube to avoid ethanol interference.)

12. Aerially pipette 50-100용출 버퍼의 μl 멤브레인 위에, 원심분리기 12,000 rpm 1 분, and collect the RNA solution.

(메모: RNA 용출 50 μl of elution buffer can increase RNA concentration but reduces total RNA yield.)