Pancreatic cancer prognosis molecular marker non-coding RNA Lnc-DCTD-25 and application thereof
A molecular marker, rnalnc-dctd-25 technology, applied in the field of tumor molecular biology, to achieve the effect of prolonging survival time, huge economic and social benefits, and improving survival rate
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Embodiment 1
[0032] Example 1 Preparation of reagents for detecting the expression of Lnc-DCTD-25 used to prepare a kit for the prognosis of patients with pancreatic cancer (for 30 reactions)
[0033] 1. Trizo l20ml
[0034] 2. Inhibit RNA degradation solvent 40ml
[0035] 3. Chloroform 80ml
[0036] 4. Isopropanol 80ml
[0037] 5. DEPC water 10ml
[0038] 6. Mixture of 10×random primers and Oligo dT primers 100μl
[0039] 7. 5× Reverse Transcription Reaction Buffer 150μl
[0040] 8. 10mM with Mg 2+100 μl of deoxyribonucleotide triphosphate dNTP
[0041] 9. 200U / μl M-MLV reverse transcriptase 50μl
[0042] 10. SYBR Green qPCR Mix 500μl
[0043] 11. 3μM target gene Lnc-DCTD-25 specific primer (its sequence is shown in SEQ NO: 2, 3) 50μl
[0044] 12. 3μM internal reference gene TUBA1A-specific primer (its sequence is shown in SEQ NO: 4, 5) 50μl
Embodiment 2
[0045] Example 2 Detection of Lnc-DCTD-25 in Tissue Samples
[0046] 1. Collect the pancreatic cancer or normal control tissues to be tested, wash them with normal saline, put them into a cryopreservation tube filled with a solvent that inhibits RNA degradation, and put them in a -80°C refrigerator for later use.
[0047] 2. Extraction of RNA in tissues:
[0048] (1) Add liquid nitrogen to the mortar first, then cut the tissue into small pieces and grind it into powder in liquid nitrogen, take 100mg of tissue powder with a liquid nitrogen pre-cooled spoon and add it to the EP tube filled with 1ml of Trizol solution middle. The total volume of tissue powder should not exceed 10% of the volume of Trizol used, and it should be thoroughly mixed evenly.
[0049] (2) Leave at room temperature for 5 minutes, then add 200 μL of chloroform, tightly cap the EP tube and shake vigorously for 0.5 minutes. Centrifuge at 12,000 rpm for 10 minutes.
[0050] (3) Take the upper aqueous phas...
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