The open reading frame of Spinacia oleracea Betaine Aldehyde Dehydrogenase (SoBADH) was retrieved from Spinacia oleracea and inserted into the Agrobacterium tumefaciens binary vector pBin438, which was driven by CaMV35S promoter, and produced the new binary vector pBSB. A. tumefaciens LBA4404 carrying this plasmid was used in genetic transformation of plants. Forty-five primary transgenic plants were detected by PCR and verified by the Southern blotting from 65 regenerated plants, of which 27 transgenic plants had only one copy of T-DNA. The Northern blotting and Western blotting analysis indicated that the SoBADH gene had been transcribed mRNA and expression protein in the transgenic cotton lines. The testing of SoBADH activity of transgenic plant leaves showed that the enzyme activity was much higher than that of the non-transgenic cotton. The growth of transgenic plants was well under the salinity and freezing stress, whereas the non-transgenic plant grew poorly and even died. Challenging with salinity, the height and fresh weight of transgenic plants was higher compared with those of non-transgenic plants. Under the freezing stress, the relative conductivity of leaf electrolyte leakage of the transgenic cotton lines was lower than that of non-transgenic plants. These results demonstrated that the SoBADH gene could over express in the exogenous plants, and could be used in genetic engineering for cotton stress resistance.
Adaptation, Physiological ; Betaine-Aldehyde Dehydrogenase ; biosynthesis ; genetics ; Cold Temperature ; Gossypium ; enzymology ; genetics ; Plants, Genetically Modified ; enzymology ; genetics ; Salinity ; Spinacia oleracea ; enzymology ; genetics ; Stress, Physiological ; genetics

In this study, the 5' -flanking proximal region of stress-induced gene encoding betaine aldehyde dehydrogenase was isolated by Adaptor-PCR and TAIL-PCR from halophyte Suaeda liaotungensis. 1993 bp sequence was obtained by sequencing. The transcription start site, which localized at 62 bases upstream of the start ATG, was predicted using TSSP-TCM program. The functional elements were analysed by PLACE programm. The SlBADH gene promoter contains the basic elements: TATA-box, CAAT-box, and stress-induced elements: salt responsed element, cold, dehydration, ABA and frozen responsed elements, WUN responsed elements and HSE. Obtaining the promoter of betaine aldehyde dehydrogenase gene from Suaeda liaotungensis provides a foundation for analyzing the stress-induced promoter elements, studying the relationship between structure and founction of the promoter, and investigating the molecular mechanism of BADH gene regulation.
Base Sequence ; Betaine-Aldehyde Dehydrogenase ; genetics ; Chenopodiaceae ; enzymology ; genetics ; Cold Temperature ; Gene Expression Regulation, Enzymologic ; Gene Expression Regulation, Plant ; Genes, Plant ; Molecular Sequence Data ; Plant Proteins ; genetics ; Promoter Regions, Genetic ; Salts ; Sequence Analysis, DNA ; Transcription Initiation Site

Plant betaine aldehyde dehydrogenase (BADH) is a physiologically important enzyme in response to salt or drought stress. In this study, two BADH genes (PeBADH1 and PeBADH2) were cloned from Populus euphratica. Both PeBADH1 and PeBADH2 genes encode the proteins of 503 amino acid residues, with a calculated molecular mass of 54.93 kDa and 54.90 kDa, respectively. Reverse transcription PCR showed the divergence of expression pattern between the PeBADH1 and PeBADH2 genes in P. euphratica. The recombinant PeBADH1 and PeBADH2 proteins were overexpressed in E. coli, and purified by Ni-affinity chromatography. The PeBADH2 protein had 1.5-fold higher enzymatic activity towards the substrate aldehyde than PeBADH1 protein. The PeBADH1 protein revealed higher thermal stability than PeBADH2 protein. These results indicated obvious functional divergence between the PeBADH1 and PeBADH2 genes.
Amino Acid Sequence ; Betaine-Aldehyde Dehydrogenase ; biosynthesis ; genetics ; Cloning, Molecular ; Escherichia coli ; genetics ; metabolism ; Gene Expression Regulation, Plant ; physiology ; Molecular Sequence Data ; Plant Proteins ; biosynthesis ; chemistry ; genetics ; Populus ; genetics ; Protein Isoforms ; chemistry ; metabolism ; Recombinant Proteins ; biosynthesis ; genetics ; isolation & purification ; Substrate Specificity