SENO Sperm Sample DNA Extraction Kit

1. Komponente kompleta reagensa

Tehnički podaci	50T	100T
Mačka. Ne.	SN0253	SN0254
Stupovi za ekstrakciju DNK (set)	50 (set)	100 (set)
Reagent Buffer SP	20 ml	2 × 20 ml
Pufer reagensa c	30 ml	2 × 30ml
Pufer za pranje 1	15 ml	2 × 15 ml
Elucijski pufer	20 ml	20 ml
Proteinaza K	1ml	2x1ml
RNaseA	1ml	2x1ml
Priručnik za upute	1	1

2. Skladištenje

Ovaj se komplet reagensa treba čuvati na sobnoj temperaturi (15-25℃) I u suhim uvjetima, s rokom trajanja 12 mjeseca. Stupovi za pročišćavanje DNK mogu se pohraniti za 1 godina u hladnom i suhom okruženju. Proteinase K and RNaza A contain preservatives, omogućavajući prijevoz na sobnoj temperaturi, Ali za dugoročno skladištenje, Treba ih držati na -20 ℃.

3. Upute za korištenje kompleta reagensa

3.1 Ovaj komplet reagensa namijenjen je istraživanju molekularne biologije i ne smije se koristiti za dijagnozu ili liječenje bolesti.

3.2 Neke komponente u kompletu reagensa sadrže iritante. Preporučuju se zaštitne mjere poput nošenja zaštitne odjeće i naočala.

3.3 Tijekom upotrebe ovog kompleta reagensa, brza centrifuga, vodena kupelji (metalna kupka), vrtlog miksera, bezvodni etanol, sterilna deionizirana voda, i EP epruvete treba pripremiti korisnik.

4. Uvod u komplet reagensa

The sperm sample genomic DNA extraction kit provides a rapid and efficient purification solution for sperm sample DNA. It achieves sample DNA purification effectively through a dedicated extraction buffer system combined with specific nucleic acid purification columns.

This kit can extract sperm sample DNA within 30 minuta, and the entire purification process does not require toxic reagents such as phenol-chloroform. The extracted DNA can be directly used for PCR, Južni blot, and other applications.

5. Eksperimentalni principi i postupci

6. Ekstrakcijski postupak

Prije početka eksperimenta:

A.Reagent Buffer SP:This buffer should be stored long-term in an environment between 2℃ and 8℃

B.Pufer reagensa c može se taložiti u uvjetima niske temperature. It is recommended to heat at 65℃ for 5 minuta. Nakon što se talog otopi, Može se normalno koristiti.

C.Pufer za pranje 1: Prije upotrebe, add the specified amount of anhydrous ethanol as indicated on the reagent bottle label. Označite ček na etiketi kako biste naznačili dodavanje bezvodnog etanola.

D. Buffer za eluciju je a 0.1X TE otopina containing a minimal amount of EDTA. If EDTA has an impact on subsequent experiments, it is recommended to use sterile deionized water as a substitute for the elution buffer.

1. Sample Processing:

Pipeta 200μl of frozen sperm, incubate at 75℃ for 10 minutes until the sperm sample is not viscous. Centrifuga na 12000 RPM za 5 minuta, aspiraj supernatantu koliko god je to moguće, and the sperm cells will sediment at the bottom of the tube.

2. Add to the sediment from the previous step: 400μl Reagent Buffer SP, 20μl Proteinase K (10 mg/ml), 20μl RNaseA (10 mg/ml).Digest at 65℃ for 10 minuta, invert and mix 6-7 times during digestion until the sample is fully digested.
3. Add an equal volume of Pufer reagensa c and an equal volume of anhydrous ethanol, and mix well by pipetting.

(For example: If you add 400μl of Reagent Buffer IV, you will need to add approximately 460μl of Reagent Buffer C and 460μl of anhydrous ethanol. A small amount of precipitation may form after adding Reagent Buffer C, but it does not affect subsequent experiments.)

4. Transfer the obtained liquid to a DNA extraction purification column (cassette) (approximately 650-700μl each time). Let it stand at room temperature for 2 minuta, centrifuge at over 8,000 RPM za 1 minuta, discard the collected waste liquid, and re-insert the collection tube into the purification column for the next step.
5. Repeat step 4, adding the remaining liquid to the DNA extraction purification column (cassette), centrifuge at over 8,000 RPM za 1 minuta, discard the waste liquid and the collection tube.
6. Postavite stupac za pročišćavanje ekstrakcije DNK (cassette) into the collection tube, dodati 300 μl pufera za pranje 1, centrifuge at over 8,000 RPM za 1 minuta, discard the waste liquid, and place the DNA extraction purification column (cassette) back into the tube for the next step.

(Bilješka: Confirm that anhydrous ethanol has been added to Wash Buffer 1.)

7. Dodati 500μl pufera za pranje 1 na stupac za pročišćavanje DNK (cassette), centrifuga na 14,000 rpm (20,000×g) za 2 minuta, extend the centrifugation time appropriately to dry the membrane more thoroughly.
8. Postavite stupac za pročišćavanje ekstrakcije DNK (cassette) u novu cijev za centrifugu, open the lid, incubate at 65℃for 2 minuta. Extend this step if necessary to evaporate ethanol as much as possible, preventing ethanol residue from affecting downstream experiments.
9. Drop-suspend 50-100μl pufera za eluciju onto the membrane, centrifuga na 12,000 RPM za 2 minuta.

(Bilješka: 1. Using 50μl of Elution Buffer to elute DNA can increase DNA concentration but decreases the overall DNA yield. 2. The eluted DNA can be reapplied to the DNA extraction purification column, centrifuged at 12,000 RPM za 2 minutes again, to increase DNA yield.)