Identification of Cd-Responsive Genes of Solanum nigrum Seedlings Through Differential Display

Abstract

Solanum nigrum is known as a Cd-hyperaccumulator: Its ability to accumulate large amounts of cadmium in leaves affords its designation as an effective phytoremediator. To identify Cd-responsive genes in S. nigrum, a nonradioactive differential display reverse transcription polymerase chain reaction technique was applied to isolate of genes whose transcription was altered in seedlings under Cd stress. A total of 48 DD bands were identified; from these bands, fragments corresponding to seven cDNAs were cloned. Reverse Northern dot-blot analysis confirmed the different expression patterns of these genes under cadmium toxicity. The homology analysis revealed that five of these cDNAs had a clear identity to Solanum species, and putative functions were assigned, including calmodulin, ascorbate peroxidase, catalase, glutamylcysteine synthetase, and ironregulated transporter. Finally, the involvement of these genes in heavy metal tolerance is discussed.

Suppressed Expression of Ascorbate Oxidase Gene Promotes Ascorbic Acid Accumulation in Tomato Fruit

“…Regulation of APX at the transcript level affects plant tolerance to drought stress (Fotopoulos et al 2008). APX gene transcription was induced in plants by various abiotic stresses including drought, salt, salt-alkaline, CdCl 2 , and abscisic acid (Xu et al 2009;Gao et al 2010). In this study, transgenic plants with AO suppression has less leaf damage and improved photosynthesis ability compared to untransformed control after drought stress treatment (Table 1).…”

Physiological adaptations to osmotic stress and characterization of a polyethylene glycol-responsive gene in Braya humilis

“…The seeds were surface-sterilized with a sodium hypochlorite solution (5%, v/v) for 8 min, washed with sterile water three times, and then sown in petri dishes containing Murashige and Skoog medium solidified with 0.8% (w/v) agar [29]. The entire process was conducted under sterile conditions.…”

“…The resulting cDNAs were synthesized using a reverse transcription system kit (Thermofisher Scientific, Waltham, MA, USA). Three 3'-oligo (dT) anchored primers (AP1-3) and seven 5'-arbitrary primers (RP1-7), described previously by Xu et al [29], were used for PCR amplification. Each sample contained 12.5 μL of Taq plus master mix (Tiangen, Beijing, China), 9.9 μL of double-distilled water, 0.5 μL of anchored primer (10 ng/μL), 0.5 μL of arbitrary primers (10 ng/μL), and 1.6 μL of template DNA.…”

“…PCR products were electrophoresed in a 1% agarose gel, stained with ethidium bromide, and visualized under ultraviolet light. All samples were analyzed in duplicate to avoid false positives in differential display banding patterns [29]. Using a gel extraction kit (Omega Biotech, Norcross, GA, USA), differentially expressed cDNA bands were excised from the agarose gel and were purified and ligated into the pGEM-T Easy vector (Promega Corp, Madison, WI, USA).…”

“…A differential display cDNA fragment was amplified from B. humilis leaves treated with 30% PEG-6000 for 4 h using a 5'-arbitrary primer (RP4, AAGAGCCCGT) and a 3'-oligo (dT) primer (AP2, AAGCTTTTTTTTTTC) at an annealing temperature of 36°C [29]. The fragment identified was approximately 1600 bp in length, and it was not present in the control or 20% PEG/4 h samples that were also analyzed (Fig.…”