Figure 3

Scanning of diverse normal human tissues, various human cell lines, and chimpanzee blood for Alu -exonization in the TIF-IA genes. (A) Low level of L2-AEx inclusion in normal tissues. cDNA from normal human tissues and cell lines was amplified by PCR analysis with primers directed to exons 1 and 2 of all three loci of TIF-IA genes. No evidence for Alu exonization was observed in chimpanzee blood. Splicing products were separated on 1.5% agarose gel. The three mRNA isoforms are shown on the right; selected PCR products were eluted and sequenced. 293T are epithelial kidney cells. BJ-1 is normal human fibroblast cell line. HeLa are epithelial cervix cells with adenocarcinoma. HT 1080 originated from a fibrosarcoma. MCF-7 is a breast-cancer cell line. P69 was derived by immortalization of human primary prostate epithelial cells. SK MEL-28 is a melanoma cell line and U2OS is an osteosarcoma cell line. (B) Splicing patterns of L2-AEx in various human leukemia cell lines. Total cytoplasmic RNA was extracted from the indicated cell lines. Splicing products were separated on 1.5% agarose gel after RT-PCR analysis, using primers to exons 1 and 2 of all three loci of TIF-IA, to locus 15 alone, to locus 28 alone, and to locus 21 alone. The three mRNA isoforms are shown on the right. Selected splicing products were eluted from the gel and sequenced. Jurkat, Molt-3, and HSB are human T-cell leukemia cell lines. HL-60 is a myeloid cell line. Dami is a megakaryocytic AMKL (acute megakaryoblastic leukemia) non-Down syndrome (DS) cell line. U937 is derived from a human histiocyic lymphoma. MT-4 is a human T-cell lymphoblast line. Dg-75, Raji, and Daudi are Burkitt's lymphoma cell lines. KM H2 is a human Hodgkin's lymphoma cell line. CMK is a megakaryocytic DS. K562 is a chronic myeloid leukemia (CMK) cell line. MEG-01 is a megakaryoblastic cell line. 697 is pre-B ALL cell line. Nalm-6 and REH are human precursor leukemia cell lines.