羊草自交不亲和性相关基因TrxLc的克隆、表达和酶活性分析

羊草是一种重要的禾本科牧草，主要分布于远东地区。硫氧化还原蛋白H (Trx h) 是一大类广泛分布于动物、植物和微生物中的小分子蛋白。在一些禾本科植物中，一种在成熟花粉中表达的Trx h，与植物的自交不亲和性有关。在本研究中, 我们克隆了羊草花粉硫氧还蛋白基因(TrxLc)的DNA和cDNA序列, 与其他禾本科植物Trx h的cDNA序列相

Molecular Cloning,Expression and Activity Assay of Thioredoxin h - A Gene Related to Self-incompatibility in Leymus chinensis

Leymus chinensis (Trin.) Tzvel., which is widely distributed in East Asia, is an important self incompatible forage species in Poaceae. h type thioredoxins are a family of small proteins that exists throughout the animal, plant, and bacterial kingdoms. Gene for thioredoxin h expressed in mature pollens is related to self incompatibility in some grasses species. In the present study, DNA and cDNA sequences of thioredoxin h ( TrxLc ) gene were cloned from L. chinensis . The cDNA nucleotide sequence of TrxLc had 53.5%-56 4% of homology to thioredoxin h genes that originated from other self incompatible grasses. Using Southern blotting, gene TrxLc was estimated to be one copy in L. chinensis genome. Northern blotting indicated that TrxLc did not express in leaves, roots, stems, and young pistils, but expressed in young pollens and mature pistils. The highest expression level of TrxLc was detected in mature pollens. Gene TrxLc was expressed as recombinant protein in Escherichia coli and tested for thioredoxin activity. In the nonspecific insulin reduction assay, the protein displayed typical thioredoxin activity and had similar values of V max and K m with h type thioredoxins of other plant species, but was not similar with that of E. coli.