Robert L S, Thompson R D, Flavell R B
AFRC Institute of Plant Science Research, Cambridge Laboratory, Trumpington, United Kingdom.
Plant Cell. 1989 Jun;1(6):569-78. doi: 10.1105/tpc.1.6.569.
The expression of a wheat genomic clone containing the entire coding sequence of the high molecular weight glutenin subunit 12 gene flanked by 2.6 kilobases of 5' and 1.5 kilobases of 3' sequences has been studied after introduction into tobacco. Seeds of different tobacco plants containing the full-length wheat genomic clone accumulated different amounts of intact high molecular weight glutenin subunit mRNA and of a polypeptide displaying the solubility, molecular weight, and antigenic properties of the high molecular weight glutenin subunit 12. The wheat protein accumulated without obvious degradation products and constituted up to approximately 0.1% of the total tobacco endosperm protein. Restriction fragments corresponding to 2.6 kilobases, 1.4 kilobases, and 433 base pairs of high molecular weight glutenin 5' upstream sequence were fused to the coding sequence of the chloramphenicol acetyltransferase (CAT) gene in the vector polyCATter and transferred into tobacco. Chloramphenicol acetyltransferase enzyme activity was detected only in the seed endosperm tissue of the transformed plants. It was detected in tobacco seeds 8 days after anthesis and persisted until seed maturity. It is concluded that 433 base pairs of high molecular weight glutenin upstream sequence are sufficient to confer endosperm-specific expression of this monocot gene in the dicot tobacco.
将一个包含高分子量谷蛋白亚基12基因完整编码序列且两侧分别有2.6千碱基的5'序列和1.5千碱基的3'序列的小麦基因组克隆导入烟草后,对其表达情况进行了研究。不同烟草植株的种子含有全长小麦基因组克隆,积累了不同量的完整高分子量谷蛋白亚基mRNA以及一种具有高分子量谷蛋白亚基12的溶解性、分子量和抗原特性的多肽。小麦蛋白积累时没有明显的降解产物,约占烟草胚乳总蛋白的0.1%。将对应于高分子量谷蛋白5'上游序列2.6千碱基、1.4千碱基和433碱基对的限制性片段与载体polyCATter中氯霉素乙酰转移酶(CAT)基因的编码序列融合,并转入烟草。仅在转化植株种子的胚乳组织中检测到氯霉素乙酰转移酶活性。在开花后8天的烟草种子中检测到该活性,并持续到种子成熟。得出的结论是,高分子量谷蛋白上游序列的433碱基对足以使该单子叶基因在双子叶烟草中进行胚乳特异性表达。