The Expression Of Metallothionein Gene Under Tissue-Specific Promoter In Rice And The Tolerance To Heavy Metals In Transgenic Lines

Mammal Metallothioneins are a family of low molecular weight metal binding proteins with a high cysteine content. MT functionally acts as detoxification of heavy metals.We have obtained transgenic rice plants containing a mouse metallothionein(mMT) gene fused to the tissue-specific promoter and constitutive promoter(35S). The transgenic rice plants were generated by transforming rice callus with Agrobacterium tumefaciens stain carrying the chimaeric gene . Transformants were directly setected by antibiotic. The existence, stable integration and inheritance to the mMT gene in T₁ generation plants were verified by Southern blot. Northern blot and Western blot indicated that the mMT gene was expressed in different tissue of transgenic rice. mMT gene fused to the constitutive promoter(CaMV 35S)was expressed in leaf and rootand that fused to the expression of tissue-specific promoter was only expressed in leaf.To study the function rice of rice metallothioneins(rMT) with the tolerance to heavy metals,we cloned three rice MTs into the RNAi vector pJawoh18RNAi which had herbicide(PPT) resistance gene bar as a selectable marker, and named pJRrMT1a﹑pJRrMT2a﹑pJRrMT3a and pJRrMT123a, The transgenic rice plants were generated by transforming rice callus with Agrobacterium tumefaciens stain carrying the chimaeric gene . Transformants were directly setected by antibiotic. The existence to the rice MT gene in T₀ and T₁ generation plants was verified by PCR and Southern blot , and the results also demonstrated the stable integration and inheritance of exotic genes. We use rice MT sequence to The level of rice MT gene expression was directly measured by RT-PCR,using ACT1 as a normalizing gene, and the results indicated that the rice MT messenger RNA was partly degraded .T1 seed were germinated in water containing antibiotic. The antibiotic tolerance trait followed Mendelian inheritance and co-segregated. T₁ generation plants with mMT gene were significantly different from the control plants with respect to Cd²⁺ intolerance. Growth of transgenic rice was unaffected by Cd²⁺ even with concentration as high as 1.0 mmol/L in the water, whereas that of control plants was severely hampered in the water containing only...