Rabbitts T H, Jarvis J M, Milstein C
Cell. 1975 Sep;6(1):5-12. doi: 10.1016/0092-8674(75)90067-7.
32P-labeled light chain messenger RNA was prepared from mouse MOPC 21 myeloma cells. The messenger RNA was hybridized to purified repetitive nuclear DNA and both the hybridized (repetitive 32P-RNA) and nonhybridized (nonrepetitive 32P-RNA) fractions were isolated. Only the nonhybridized RNA gave a T1 ribonuclease fingerprint showing oligonucleotides derived from the variable and constant regions of the light chain messenger RNA. In addition, this fingerprint showed oligonucleotides derived from the untranslated regions of the light chain messenger RNA. The nonrepetitive 32P-RNA was shown to rehybridize only with the unique fraction of total nuclear DNA. The rapidly hybridizing part of the unfractionated 32P-RNA preparation, therefore, is not a component of the light chain messenger RNA itself. Complementary DNA was prepared with reverse transcriptase using unlabeled light chain messenger RNA as template, and the transcripts were fractionated into various size classes. Complementary DNA molecules greater than 900 bases in length hybridized with both the initial messenger RNA and with the nonrepetitive 32P-RNA but failed to hybridize with excess purified repetitive 32P-RNA. The rapidly hybridizing component of the messenger RNA fraction, therefore, does not appear to be transcribed by reverse transcriptase. It is concluded that, under the experimental conditions used, the light chain messenger RNA hybridizes exclusively with unique DNA.
从鼠MOPC 21骨髓瘤细胞制备了32P标记的轻链信使核糖核酸。将该信使核糖核酸与纯化的重复性核DNA杂交，并分离出杂交部分（重复性32P-RNA）和未杂交部分（非重复性32P-RNA）。只有未杂交的RNA产生了T1核糖核酸酶指纹图谱，显示出源自轻链信使核糖核酸可变区和恒定区的寡核苷酸。此外，该指纹图谱还显示出源自轻链信使核糖核酸非翻译区的寡核苷酸。已证明非重复性32P-RNA仅与总核DNA的单一部分重新杂交。因此，未分级的32P-RNA制剂中快速杂交的部分不是轻链信使核糖核酸本身的组成部分。以未标记的轻链信使核糖核酸为模板，用逆转录酶制备互补DNA，并将转录本分级为不同大小类别。长度大于900个碱基的互补DNA分子与初始信使核糖核酸和非重复性32P-RNA都杂交，但不能与过量纯化的重复性32P-RNA杂交。因此，信使核糖核酸部分中快速杂交的成分似乎不是由逆转录酶转录的。得出的结论是，在所使用的实验条件下，轻链信使核糖核酸仅与单一DNA杂交。
