Ly-6E.1 htal-1 and Ly-6E.1 Δbhtal-1 vectors and transgene expression.

(A) The Ly-6E.1 vector contains the 14-Kb Ly-6E.1genomic sequence, including promoter and 3′ sequence of theLy-6E.1 gene. A human tal-1(htal-1) cDNA or Δbhtal-1 cDNA were linked to the bovine growth hormone polyA sequence and inserted in the first exon of the Ly-6E.1 gene as previously described.18The ΔbTAL-1 protein lacks the basic domain-containing amino acids 187 to 199.22 (B) Photo of wild-type (Wt; left) and a representative high-copy transgenic adult mouse (Tg; right) showing the shorter length and kinked tail of the transgenic mice. (C) Expression of htal-1 mRNA in a transgenic L6 mouse. Northern blot analysis of 15 μg total RNA isolated from different tissues was hybridized with a human tal-1 cDNA probe. The length of the transgene transcript is 1.2 Kb. (D) Expression of human TAL-1 protein in high copy number L6 transgenic mice. Total protein extracts of different tissues of transgenic or nontransgenic mice were separated on a 12% sodium dodecyl sulfate–polyacrylamide gel electrophoresis, transferred to Hybond-P polyvinylidene fluoride membrane (Amersham) and probed with BTL-136 that recognizes the human and mouse TAL-1 protein. Nuclear extracts from Jurkat cells were used as a positive control. The 42-kd hTAL-1 protein is indicated by an arrow. (E) Level of TAL-1 protein expression in animal lines L6, L8, and wild-type littermates. Total splenic protein extracts (80 μg) of the 10 copy L6, 2 copy L8, and wild-type animal lines were subjected to Western blot analysis using the human TAL-1 protein-specific antibody BTL-73 monoclonal antibody. Quantification of hTAL-1 protein shows that its level in L6 splenocytes is 5 times the level of L8 splenocytes. No TAL-1 protein is detected in wild-type controls.