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. 2011 Nov 14;287(4):2500–2508. doi: 10.1074/jbc.M111.303123

FIGURE 2.

FIGURE 2.

Characterization of 70Z/3-KD and 70Z/3-KD-re cells. A, gene-silencing effects of siRNA on the Fut8 mRNA expression were determined by real-time PCR, and normalized by the levels of GAPDH (n = 3). B, analyses of Fut8 activity. Fut8 activity was examined using a fluorescence-labeled sugar chain, GnGn-Asn-PABA (4-(2-pyridylamino)butylamine), as an acceptor substrate, as described under “Experimental Procedures.” The substrate (S) and Fut8 product (P) were eluted at 10 and 20 min, respectively, in mock and restored cells but not in 70Z/3-KD cells. ND, not detectable. Restored, re-introduction of Fut8 gene into 70Z/3-KD cells. C, gene expressions of Fut8, GnTIII, and β4GalT-I by real-time PCR analysis. RNAs were isolated from 70Z/3 and 70Z/3-KD cells. All values were normalized to that of the GAPDH gene. 70Z/3-KD cell, Fut8 knockdown 70Z/3 cell; 70Z/3-KD-re cell, Fut8 restored 70Z/3-KD cells. Data were representative of the mean ± S.D. of 3 per genotype (**, p < 0.01).