Application of VGLL4 gene in treatment of tumors and related medicament of VGLL4 gene
A gene and drug technology, applied in the biological field, can solve problems such as excessive cell proliferation and increased risk of cancer
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Embodiment 1
[0091] Example 1 The relationship between the expression of VGLL4 and the occurrence and development of gastric cancer
[0092] 1. Experimental method
[0093] In order to study the clinical relationship between VGLL4 and YAP, the mRNA levels of VGLL4 and YAP were detected in 84 cases of human gastric cancer samples and 91 cases of normal human gastric tissue samples. The total RNA was extracted with guanidine isothiocyanate in one step, the first strand was synthesized by AMV reverse transcriptase, and PCR amplification was performed with the corresponding primers under the action of DNA taq enzyme using AMV reverse transcriptase; see Table 1 for the PCR primers, Realtime - See Table 2 for the PCR reaction system, see Table 3 for the Realtime-PCR amplification program, and see Table 3 for the PCR amplification results figure 1 A and figure 1 b.
[0094] Table 1 Primer Sequence
[0095]
[0096]Table 2 Realtime-PCR reaction system
[0097]
[0098] Table 3 PCR react...
Embodiment 2
[0106] Example 2 VGLL4 inhibits YAP-mediated TEAD4 transcriptional activation and cell proliferation
[0107] 1. Purpose of the experiment
[0108] The purpose of this example is to study the regulation mode among VGLL4, YAP and TEAD4 and their influence on cell proliferation.
[0109] 2. Experimental method
[0110] 2.1 Plasmid construction
[0111] 1) Construction of pCDH-YAP plasmid and pCDH-VGLL4 plasmid: One-step extraction of total RNA from human HEK293 cells with guanidine isothiocyanate, synthesis of the first strand by AMV reverse transcriptase, and using it as a template for high-fidelity DNA polymerization in Takara Enzyme PrimeSTAR PCR amplification was performed with the corresponding primers under the action of the corresponding primers; the PCR primers are shown in Table 4, the PCR reaction system is shown in Table 5, and the PCR amplification program is shown in Table 6.
[0112] Table 4 Primer sequences
[0113]
[0114] Table 5 PCR reaction system
...
Embodiment 3
[0137]Example 3 Vgl4-TEAD4 interaction affects Vgl4 inhibition of YAP-mediated transcriptional activation of TEAD4 and its downstream genes
[0138] 1. Experimental materials
[0139] 1.1 Construction of plasmid
[0140] 1) HT-pET-28a vector: commercialized pET-28a (Novagen) was used as a template, and the following primers were used for construction. The PCR amplification program is: 98°C 2min; PCR cycle: 98°C 10s, 58°C 10s, 68°C 1kb / min, 18 cycles; 68°C 10min. Use DpnI demethylase to remove the unmutated wild-type pET28a template, transform the PCR amplified fragment into DH5α competent cells, and use the characteristic of E. coli to repair the loss of the vector to circularize the entire fragment to form a vector. After screening, the HT-pET-28a vector with the same backbone as the original pET28a vector but with TEV restriction sites and common multiple cloning sites was obtained.
[0141] HT-s:
[0142] 5'-Gaaaacctgtattttcagggcgccatggatccggaattcaaaggcctacgtcgacgagctca...
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