The polymorphisms of Waxy (Wx) microsatellite and G-T single-nucleotide polymorphism (SNP) in the Wx gene region were analyzed using simplified techniques in fifteen rice varieties. A rapid and reliable electrophoresis method, MetaPhor agarose gel electrophoresis (MAGE), was effectively employed as an alternative to polyacrylamide gel electrophoresis (PAGE) for separating Wx microsatellite alleles. The amplified products containing the Wx microsatellite ranged from 100 to 130 bp in length. Five Wx microsatellite alleles, namely (CT)10, (CT)11, (CT)16, (CT)17, and (CT)18 were identified. Of these, (CT)11 and (CT)17 were the predominant classes among the tested varieties. All varieties with an apparent amylose content higher than 24% were associated with the shorter repeat alleles; (CT)10 and (CT)11, while varieties with 24% or less amylose were associated with the longer repeat alleles. All varieties with intermediate and high amylose content had the sequence AGGTATA at the 5′-leader intron splice site, while varieties with low amylose content had the sequence AGTTATA. The G-T polymorphism was further verified by the PCR-AccI cleaved amplified polymorphic sequence (CAPS) method, in which only genotypes containing the AGGTATA sequence were cleaved by AccI. Hence, varieties with desirable amylose levels can be developed rapidly using the Wx microsatellite and G-T SNP, along with MAGE.

Background Bacillus thuringiensis (Bt) produces a group of δ-endotoxin proteins designated as cry toxins. No doubt that Bt isolates are excellent potential candidates for biological control strategies. Results The present study showed that 12 Bt strains were isolated and characterized at morphological, biochemical, and molecular levels. All the tested 12 Bt strains were gram-positive, endospore-forming, and possessing typical Bt crystal structures under the scanning electron microscopy (SEM). Universal primers direct and reverse of five pairs were used to detect five Cry-type genes (Cry1, Cry2, Cry3, Cry4, Cry7, and Cry8) by the PCR sizes produced from the studied Bt strains. The 16S rRNA PCR technique, 16S gene primer, DNA template, dNTPs, and Taq polymerase produced unique and distinguishable restriction patterns used for the molecular characterization of the studied Bt strains. Based on the PCR products, the frequency of Cry-gene distribution among the tested strains was Cry1 100%, Cry4 85%, and Cry3 62%, and Cry2 and Cry7 frequency was 54%. Based on the activity of insecticidal of the tested Bt strains, Bt1, Bt9, Bt10, and Bt11 were extremely pathogenic; their pathogenicity ranged from 93 to 100% against dipteran and lepidopteran larvae, compared to the other Bt isolates. The nucleotide sequences of amplified 1500 bp conserved region of 16S rRNA genes of four strains blasted using NCBI database compared to NCBI database sequences, and they were reported as native strains of Bt showing high homology with the known Bt strains (99–100%). The nucleotide sequences of Bt1, Bt9, Bt10, and Bt11 were placed in the GenBank database under accession numbers MN860017, MN843958, MN843959, and MN843960, respectively. Conclusion The strategies of enhancing the sustainability of crops and vegetables that are targeted by a large number of pathogenic insects require a great effort of exploring novel species and strains of Bt. Herein, native strains of Bt were documented from the eastern province of Saudi Arabia that displayed bio-insecticidal action on larvae of Diptera and Lepidoptera.

No abstract

This study aimed to determine the effects of different concentrations and combinations of the phytohormones 2,4-dichlorophenoxy acetic acid (2,4-D), kinetin, 6-benzylaminopurine (BAP), and 1-naphthaleneacetic acid (NAA) on callus induction and to demonstrate the role of elicitors and exogenous precursors on the production of mitragynine in a Mitragyna speciosa suspension culture. The best callus induction was achieved from petiole explants cultured on WPM that was supplemented with 4 mg L−1 2, 4-D (70.83%). Calli were transferred to liquid media and agitated on rotary shakers to establish Mitragyna speciosa cell suspension cultures. The optimum settled cell volume was achieved in the presence of WPM that contained 3 mg L−1 2,4-D and 3% sucrose (9.47 ± 0.4667 mL). The treatment of cultures with different concentrations of yeast extract and salicylic acid for different inoculation periods revealed that the highest mitragynine content as determined by HPLC was achieved from the culture treated with 250 mg L−1 yeast extract (9.275 ± 0.082 mg L−1) that was harvested on day 6 of culturing; salicylic acid showed low mitragynine content in all concentrations used. Tryptophan and loganin were used as exogenous precursors; the highest level of mitragynine production was achieved in cultures treated with 3 μM tryptophan and harvested at 6 days (13.226 ± 1.98 mg L−1).