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Expression of the spinach betaine aldehyde dehydrogenase (BADH) gene in transgenic tobacco plants. Chin J Biotechnol

Institute of Botany, Chinese Academy of Sciences, Beijing, China.
Chinese journal of biotechnology 02/1997; 13(3):153-9.
Source: PubMed

ABSTRACT Plasmid pLS9 contains a 1.5-kb of spinach cDNA including its complete open reading frame. The 1.5-kb BADH cDNA was cut from pLS9 using restriction enzyme and was inserted into the expression cassette of plasmid pYH between the CaMV 35S promoter and polyA signal sequence. The 35S-BADH cDNA-polyA fragment of pYH was cloned into a polylinker cloning site of the binary vector pBin19. The resulting plasmid pBinBADH-S was transferred to Agrobacterium tumefacies LBA4404. The tobacco plants were transformed with strain LBA4404 containing pBinBADH-S, and more than ninety kanamycin-resistant transformants were selected. Polymerase chain reaction (PCR) detection showed that more than 60% of the transformed tobacco plants contained the foreign BADH gene. The Western blot analysis, BADH enzymatic assay, specific stain for BADH activity, and the test for salt tolerance showed that BADH gene was normally expressed in the transgenic tobacco plants. The BADH enzymes also presented in chloroplasts and cytosol of the transgenic plants. The transgenic tobacco plants having strong expression of BADH gene had strong ability to tolerate high salt stress.

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    • "mangrove , spinach, amaranth, barley and sorghum, are proven betaine accumulators and tolerate salt and drought stress partly through this mechanism, but other species like tobacco, tomato and rice are considered non-accumulators of GB (Ishitani et al. 1993; Rathinasabapathi et al. 1993; Shirasawa et al. 2006). Transformation of the Badh gene from bacterial and plant sources into betaine-deficient plant species has resulted in accumulation of GB in their system and consequent acquisition of tolerance to salt and drought stress (Liang et al. 1997; Mohanty et al. 2002). BADH synthesis is up-regulated several-fold in response to salt and drought stress in spinach, barley and sorghum leaves (Weretilnyk and Hanson 1990; Ishitani et al. 1995; Wood et al.1996). "
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    ABSTRACT: Betaine aldehyde dehydrogenase (BADH) is a key enzyme involved in the synthesis of glycinebetaine—a powerful osmoprotectant against salt and drought stress in a large number of species. Rice is not known to accumulate glycinebetaine but it has two functional genes coding for the BADH enzyme. A non-functional allele of the BADH2 gene located on chromosome 8 is a major factor associated with rice aroma. However, similar information is not available regarding the BADH1 gene located on chromosome 4 despite the similar biochemical function of the two genes. Here we report on the discovery and validation of SNPs in the BADH1 gene by re-sequencing of diverse rice varieties differing in aroma and salt tolerance. There were 17 SNPs in introns with an average density of one per 171 bp, but only three SNPs in exons at a density of one per 505 bp. Each of the three exonic SNPs led to changes in amino acids with functional significance. Multiplex SNP assays were used for genotyping of 127 diverse rice varieties and landraces. In total 15 SNP haplotypes were identified but only four of these, corresponding to two protein haplotypes, were common, representing more than 85% of the cultivars. Determination of population structure using 54 random SNPs classified the varieties into two groups broadly corresponding to indica and japonica cultivar groups, aromatic varieties clustering with the japonica group. There was no association between salt tolerance and the common BADH1 haplotypes, but aromatic varieties showed specific association with a BADH1 protein haplotype (PH2) having lysine144 to asparagine144 and lysine345 to glutamine345 substitutions. Protein modeling and ligand docking studies show that these two substitutions lead to reduction in the substrate binding capacity of the BADH1 enzyme towards gamma-aminobutyraldehyde (GABald), which is a precursor of the major aroma compound 2-acetyl-1-pyrroline (2-AP). This association requires further validation in segregating populations for potential utilization in the rice breeding programs.
    Molecular Breeding 03/2010; 26(2). DOI:10.1007/s11032-010-9425-1 · 2.28 Impact Factor
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    • "Bradbury et al. (2008) reported that the OsBADH1 and OsBADH2 enzymes are substrate-specific; for example, OsBADH2 have higher specificity to betaine aldehyde, which is the intermediate substrate for the glycine betaine biosynthesis pathway, than OsBADH1 enzyme. Although OsBADH2 has higher affinity to betaine aldehyde than OsBADH1, OsBADH1 has been reported to exhibit significantly increased transcriptional level when exposed to salt and drought stresses (Niu et al. 2007; Ishitani et al. 1994; Liang et al. 1997), whereas no consistent relationship between OsBADH2 transcription level and salt treatment was observed (Fitzgerald et al. 2008). This evidence suggests that only the OsBADH1 gene is probably involved in protecting plants against these stresses. "
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    ABSTRACT: Glycine betaine has been reported as an osmoprotectant compound conferring tolerance to salinity and osmotic stresses in plants. We previously found that the expression of betaine aldehyde dehydrogenase 1 gene (OsBADH1), encoding a key enzyme for glycine betaine biosynthesis pathway, showed close correlation with salt tolerance of rice. In this study, the expression of the OsBADH1 gene in transgenic tobacco was investigated in response to salt stress using a transgenic approach. Transgenic tobacco plants expressing the OsBADH1 gene were generated under the control of a promoter from the maize ubiquitin gene. Three homozygous lines of T2 progenies with single transgene insert were chosen for gene expression analysis. RT-PCR and western blot analysis results indicated that the OsBADH1 gene was effectively expressed in transgenic tobacco leading to the accumulation of glycine betaine. Transgenic lines demonstrated normal seed germination and morphology, and normal growth rates of seedlings under salt stress conditions. These results suggest that the OsBADH1 gene could be an excellent candidate for producing plants with osmotic stress tolerance.
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    • "Nicotiana tabacum cv. Honghuadajinyuan was used as plant materials and was transformed by the standard Agrobacterium-mediated method (Liang et al. 1997). Transformants (T 0 ) were selected on MS agar containing 50 mg l -1 hygromycin. "
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    ABSTRACT: Aldehyde dehydrogenases (ALDHs) play a central role in detoxification processes of aldehydes generated in plants when exposed to the stressed conditions. In order to identify genes required for the stresses responses in the grass crop Zea mays, an ALDH (ZmALDH22A1) gene was isolated and characterized. ZmALDH22A1 belongs to the family ALDH22 that is currently known only in plants. The ZmALDH22A1 encodes a protein of 593 amino acids that shares high identity with the orthologs from Saccharum officinarum (95%), Oryza sativa (89%), Triticum aestivum (87%) and Arabidopsis thaliana (77%), respectively. Real-time PCR analysis indicates that ZmALDH22A1 is expressed differentially in different tissues. Various elevated levels of ZmALDH22A1 expression have been detected when the seedling roots exposed to abiotic stresses including dehydration, high salinity and abscisic acid (ABA). Tomato stable transformation of construct expressing the ZmALDH22A1 signal peptide fused with yellow fluorescent protein (YFP) driven by the CaMV35S-promoter reveals that the fusion protein is targeted to plastid. Transgenic tobacco plants overexpressing ZmALDH22A1 shows elevated stresses tolerance. Stresses tolerance in transgenic plants is accompanied by a reduction of malondialdehyde (MDA) derived from cellular lipid peroxidation.
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