Antitumor drug screening kit and use method thereof
A technology of anti-tumor drugs and kits, which is applied in the field of biomedicine to achieve the effects of objective and accurate data, simple operation and short cycle.
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Embodiment 1
[0051] Example 1 Confirmatory Experiment of the Staining Effect of Caenorhabditis elegans
[0052] (1) Synchronization of Caenorhabditis elegans
[0053] ① Select mature Caenorhabditis elegans into a 2ml centrifuge tube containing 1ml M9 solution, centrifuge at 4000rpm for 5 minutes, and remove the supernatant;
[0054] ② Add 500 μL of 1M sodium hydroxide and 500 μL of 6.4% (V / V) sodium hypochlorite to the centrifuge tube, and shake to lyse;
[0055] ③Remove the supernatant, add 1.5μL M9 solution to the pellet, shake well, centrifuge at 4000rpm for 5 minutes, repeat this step three times;
[0056] ④ Add 500 μL of M9 solution to the washed eggs, and hatch at 18°C for 24 hours to form L1-stage Caenorhabditis elegans.
[0057] (2) Staining and detection method of Caenorhabditis elegans
[0058] ①Staining and detection of Caenorhabditis elegans
[0059] Mix the above synchronized L1 stage Caenorhabditis elegans MT2124, count the number of Caenorhabditis elegans per unit liqu...
Embodiment 2
[0067] Example 2 The effect of anti-tumor candidate drugs on Caenorhabditis elegans CB1275 (lin-1 deletion mutation)
[0068] lin-1 is a gene downstream of ras, a nuclear transcription factor that can be phosphorylated by MAPK, a negative regulator of vulva development, and can regulate the signaling pathway of let-60 downstream of vulva development. lin-1 and lin-31 form a complex, lin-1 encodes ETS family members, and lin-31 encodes a winged helix transcription factor, when the ras signal is not activated, lin-1 and lin-31 interact to form a complex, This complex binds to the promoters of the ovipositor functional genes and prevents their expression. After the lin-1 deletion mutation, the complex cannot be formed, the complex cannot be formed, and it cannot be combined with the promoter part of the ovipositor function gene, and the ovipositor gene is expressed, and then the pseudovulva develops to form a polyvulva phenotype.
[0069] If the results of Example 1 show that th...
Embodiment 3
[0071] The assembly of embodiment 3 antitumor drug screening kits
[0072] Antitumor drug screening kit of the present invention, the kit includes the following reagents:
[0073] ① Caenorhabditis elegans wild type N2;
[0074] ② Caenorhabditis elegans mutant MT2124;
[0075] ③ Caenorhabditis elegans CB1275;
[0076] ④ NGM medium;
[0077] ⑤ Escherichia coli OP50;
[0078] ⑥Dye: 2',7'-dichlorodihydrofluorescein diethyl ester.
[0079] Wherein, the concentration of Caenorhabditis elegans is 100 / 20 μl, the concentration of Escherichia coli OP50 is 10 mg / ml, and the final concentration of the dye is 50 μg / mL.
[0080] Dispense the above reagents into suitable containers and put them into an outsourcing kit, in which M9 buffer, S buffer and LB medium are described in detail in the instruction manual.
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