Partial purification and characterization of the mRNA complementing a temperature-sensitive S-phase cell cycle mutation.
E36ts24 is a temperature-sensitive (ts) cell cycle mutant derived from the Chinese hamster lung cell line E36. At the restrictive temperature for growth (40.3 degrees C), the mutant cells are arrested at early S phase. We have microinjected poly(A)+ RNA isolated from the wild-type E36 cells into the cytoplasm of E36ts24 cells arrested at early S phase after 24 hr of incubation at 40.3 degrees C. The ts mutation was transiently complemented in a significant fraction of the microinjected cells as evidenced by the incorporation of [3H]thymidine assayed by autoradiography. Microinjection of mRNA fractionated by methylmercuric hydroxide/agarose gel showed that the mRNA capable of transiently complementing the mutation in E36ts24 contains about 940 nucleotides. Hence, it can code at most for a protein containing about 230 amino acids. We estimate that the partial purification by fractionation of the mRNA active in the transient complementation is on the order of 100- to 200-fold.
D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D005816:Genetic Complementation Test; D007399:Interphase; D008168:Lung; D008970:Molecular Weight; D009154:Mutation; D011061:Poly A / Q000235:genetics / Q000302:isolation & purification; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D012333:RNA, Messenger / Q000235:genetics* / Q000302:isolation & purification; D013696:Temperature; D013937:Thymidine Kinase / Q000235:genetics
D012333:RNA, Messenger; D011061:Poly A; D012313:RNA; D013937:Thymidine Kinase