Increasing the distance between the snRNA promoter and the 3' box decreases the efficiency of snRNA 3'-end formation.
Chimeric genes which contained the mouse U1b snRNA promoter, portions of the histone H2a or globin coding regions and the U1b 3'-end followed by a histone 3'-end were constructed. The distance between the U1 promoter and the U1 3' box was varied between 146 and 670 nt. The chimeric genes were introduced into CHO cells by stable transfection or into Xenopus oocytes by microinjection. The efficiency of utilization of the U1 3' box, as measured by the relative amounts of transcripts that ended at the U1 3' box and the histone 3'-end, was dependent on the distance between the promoter and 3'-end box. U1 3'-ends were formed with >90% efficiency on transcripts shorter than 200 nt, with 50-70% efficiency on transcripts of 280-400 nt and with only 10-20% efficiency on transcripts >500 nt. Essentially identical results were obtained after stable transfection of CHO cells or after injecting the genes into Xenopus oocytes. The distance between the U1 promoter and the U1 3' box must be <280 nt for efficient transcription termination at the U1 3' box, regardless of the sequence transcribed.
Ramamurthy
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Program in Molecular Biology and Biotechnology, University of North Carolina, Chapel Hill 27599, USA.
Ingledue
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TC
Pilch
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Kay
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BK
Marzluff
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D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D006224:Cricetinae; D051379:Mice; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011401:Promoter Regions, Genetic; D012342:RNA, Small Nuclear / Q000235:genetics*; D014158:Transcription, Genetic; D014162:Transfection
D012342:RNA, Small Nuclear