Transcription of the alpha globin gene domain in normal and AEV-transformed chicken erythroblasts: Mapping of giant globin-specific RNA including embryonic and adult genes

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Abstract
The genomic domain of about 20 kbp of the chicken alpha-type globin genes, framed by AT-rich linkers (ATRLs; Moreau et al. 1982) and repetitive sequences (Broders et al. 1986), was cut into 13 fragments and subcloned. The in vitro labelled individual restriction fragments were used to test the extent of the transcribed domain by blot-hybridization of nuclear RNA in large excess from normal adult chicken and Avian Erythroblastosis Virus (AEV)-transformed erythroblasts. In both these types of cells, the AT-rich segments situated 6 kbp upstream of the first gene as well as all the domain including the embryonic pi and the adult alpha D and alpha A genes down to the AT-rich segment placed 3 kbp downstream were found to be transcribed. Electrophoresis of nuclear RNA, Northern blotting and hybridization with most of the nick-translated DNA probes revealed in all cases the presence of heterogeneous globin RNA molecules in the 3–13 kb range, as well as some distinct RNA bands. Single-stranded RNA probes of some genomic segments indicated asymmetrical transcription of the minus strand. A 12 kb globin-specific RNA including the pi and alpha A genes but not the intervening alpha D gene was observed in AEV-transformed cells: it includes sequences located far upstream and downstream from the alpha globin genes and might represent a processing product of a full length transcript spanning the whole domain. Reverse transcription by extension of primers placed in the first exon of each of the three globin genes confirmed the presence of continuous transcripts of the domain including the two adult and the embryonic globin genes.