Elhag G A, Thomas F J, McCreery T P, Bourque D P
Department of Biochemistry, University of Arizona, Tucson 85721.
Nucleic Acids Res. 1992 Feb 25;20(4):689-97. doi: 10.1093/nar/20.4.689.
Poly(A)+ mRNA isolated from Nicotiana tabacum (cv. Petite Havana) leaves was used to prepare a cDNA library in the expression vector lambda gt11. Recombinant phage containing cDNAs coding for chloroplast ribosomal protein L12 were identified and sequenced. Mature tobacco L12 protein has 44% amino acid identity with ribosomal protein L7/L12 of Escherichia coli. The longest L12 cDNA (733 nucleotides) codes for a 13,823 molecular weight polypeptide with a transit peptide of 53 amino acids and a mature protein of 133 amino acids. The transit peptide and mature protein share 43% and 79% amino acid identity, respectively, with corresponding regions of spinach chloroplast ribosomal protein L12. The predicted amino terminus of the mature protein was confirmed by partial sequence analysis of HPLC-purified tobacco chloroplast ribosomal protein L12. A single L12 mRNA of about 0.8 kb was detected by hybridization of L12 cDNA to poly(A)+ and total leaf RNA. Hybridization patterns of restriction fragments of tobacco genomic DNA probed with the L12 cDNA suggested the existence of more than one gene for ribosomal protein L12. Characterization of a second cDNA with an identical L12 coding sequence but a different 3'-noncoding sequence provided evidence that at least two L12 genes are expressed in tobacco.
从烟草（品种为 Petit Havana）叶片中分离得到的聚腺苷酸加尾（Poly(A)+）mRNA 被用于在表达载体λgt11 中构建 cDNA 文库。鉴定并测序了包含编码叶绿体核糖体蛋白 L12 的 cDNA 的重组噬菌体。成熟的烟草 L12 蛋白与大肠杆菌的核糖体蛋白 L7/L12 具有 44%的氨基酸同一性。最长的 L12 cDNA（733 个核苷酸）编码一个分子量为 13,823 的多肽，其具有一个由 53 个氨基酸组成的转运肽和一个由 133 个氨基酸组成的成熟蛋白。转运肽和成熟蛋白分别与菠菜叶绿体核糖体蛋白 L12 的相应区域具有 43%和 79%的氨基酸同一性。通过对 HPLC 纯化的烟草叶绿体核糖体蛋白 L12 进行部分序列分析，证实了成熟蛋白预测的氨基末端。通过将 L12 cDNA 与聚腺苷酸加尾（Poly(A)+）RNA 和总叶 RNA 杂交，检测到一条约 0.8 kb 的单一 L12 mRNA。用 L12 cDNA 探测烟草基因组 DNA 的限制性片段的杂交模式表明，核糖体蛋白 L12 存在不止一个基因。对第二个具有相同 L12 编码序列但不同 3'非编码序列的 cDNA 的表征提供了证据，表明烟草中至少有两个 L12 基因表达。
