Efficacy of RNA interference after a transfection of SALL4 shRNA vector in pediatric testicular yolk sac tumor

Abstract

Objective To construct the special SALL4 shRNA vector for transfection into cultured testicular yolk sac tumor cells, evaluate the effect of interference and provide experimental rationales for elucidating the function of SALL4. Methods Four special interference siRNAs and 1 negative siRNA were designed for different interference foci. And pGPU6/GFP/Neo-SALL4 shRNA vector was constructed and transfected into blank, negative(NC), MOCK and interference groups of testicular yolk sac tumor cells. The expression quantities of SALL4 mRNA and protein were measured by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and Western blot. And the optimal interference sequence was selected. Results The mRNA expression magnitudes of Blank, negative(NC), MOCK group, SALL4-homo-483, SALL4-homo-951, SALL4-homo-1874 and SALL4-homo-3110 were(100)%, (118.0±18.0)%, (128.0±15.0)%, (91.0±21.0)%, (25.0±8.0)%, (59.0±11.0)% and(49.0±13.0)% respectively. As compared with Blank, NC and MOCK, SALL4-homo-951 and SALL4-homo-3110 had lower levels of mRNA expression. And the protein expressions of Blank, NC, MOCK group, SALL4-homo-483, SALL4-homo-951, SALL4-homo-1874 and SALL4-homo-3110 were 1, 1.1±0.1, 1.3±0.3, 1.7±0.4, 0.2±0.1, 0.9±0.2 and 0.4±0.1 respectively. SALL4-homo-951 and SALL4-homo-3110 lowered the SALL4 protein expression. Conclusions SALL4-homo-951 and SALL4-homo-3110 may interfere with the expression of SALL4 in testicular yolk sac tumor cells. Especially the former has the optimal efficacy. Key words: Yolk sac tumor of testis; Gene expression; Transfection