Randomized trial examines the influence of DNA sequences on RNA production in mouse L cells, indicating critical regions for transcription initiation.
Key Points
To identify the essential DNA sequences required for correct beta-globin RNA formation in mouse L cells.
Transformed mouse thymidine kinase-negative (TK-) L cells with cloned herpes simplex virus 1 TK DNA and rabbit beta-globin DNA with various flanking sequences.
Selected TK+ cell lines to evaluate RNA production from different flanking sequence lengths of 14, 66, 76, 425, and 1500 nucleotides.
Mapped 5' termini of produced RNA transcripts to determine the effectiveness of transcription initiation.
With 14 base pairs of flanking sequence, none of the RNA transcripts had the correct 5' end; most originated from the vector.
With 66 base pairs, 5% of transcripts had correct 5' termini; with 76 or more base pairs, 30-85% were accurate.
The region between 14 and 66 base pairs is essential for initiating transcription correctly.