[Show abstract][Hide abstract] ABSTRACT: Vigna plants distributed in Korea were analyzed by molecular cytogenetic fluorescence in situ hybridization (FISH), genomic in situ hybridization (GISH) and rDNA ITS/NTS sequences. FISH revealed that variable 45S rRNA gene loci (one to four) were localized on the terminal regions of chromosomes, while two conserved 5S rRNA gene loci from all species examined, except for rice bean (single locus), were detected. FISH and GISH showed the characteristic organization of rRNA gene loci and genomic homology on the chromosomes, indicating their cytogenetic relatationships. ITS sequence revealed that there was considerable variation in length (190–207 bp in ITS1, 205–221 bp in ITS2) and nucleotide composition (7–67 bp). The 5S rRNA gene unit comprised coding region (118 bp) and extensive sequence heterogeneity (97–221 bp). Phylogenetic analysis of the ITS and NTS sequences demonstrated that the Vigna species are divided into two groups: angularis (V. angularis, V. umbellata, V. nakashimae and V. nipponensis) and unguiculata (V. unguiculata, V. sesquipedalis and V. vexillata). Sequence data also showed that mung bean was closer to the angularis group.
[Show abstract][Hide abstract] ABSTRACT: Internal transcribed spacers (ITS) of nuclear ribosomal DNA cistrons were studied in wild Lilium species native to Korea and compared with those species in the genus distributed in other countries to investigate the phylogenetic relationships among Lilium species. Sequence analysis data of Lilium plants collected from China and Japan were also included to study the inter- and intra-specific relationships in the genus. Lilium species formed a monophyletic clade and were separated in two clusters that belonged to the section Sinomartagon and Martagon. The phylogenetic relationships of Lilium species inferred from the ITS sequences using the maximum likelihood method were highly congruent with previous studies of cytogenetic evidence and classification (Comber, 1949) based on the morphological characters. The length of ITS1 and ITS2 regions in Lilium species native to Korea was uniform, however, nucleotide polymorphisms were noted. A region with a conserved sequence was observed from position 330–340 among all the species. Comparison of the sequence analysis of Lilium accessions native to different countries revealed the presence of six insertions/deletions (indels) and 121 substitutions (transitions and transversions) in both regions of ITS1 and ITS2. Indels were responsible for variation of ITS length in different accessions. The sequences of Lilium species collected from each country had less variations than those of species collected from different countries. Sequences variations in both ITS regions suggested that they could be used for assaying genetic diversity in Lilium at the inter- and intra-specific level.
Preview · Article · Oct 2011 · Horticulture, Environment and Biotechnology
[Show abstract][Hide abstract] ABSTRACT: AtATM3, a member of the ATP-binding cassette transporter family, is localized at the mitochondrial membrane of Arabidopsis thaliana and is involved in the biogenesis of Fe–S clusters and iron homeostasis in plants. Through Agrobacterium-mediated genetic transformation, the AtATM3 gene driven by the cauliflower mosaic virus 35S promoter (CaMV35S) was introduced into Brassica juncea (Indian mustard), a plant species suitable for phytoremediation, with the aim of improving heavy metal tolerance and accumulation
in plants. The presence of the AtATM3 gene in transgenic plants was confirmed by polymerase chain reaction (PCR). Reverse transcriptase-PCR analysis confirmed
AtATM3 expression in transgenic plants, but the level of AtATM3 expression varied between lines. AtATM3 overexpression in B. juncea conferred enhanced tolerance to cadmium [Cd(II)] and lead [Pb(II)] stresses. Importantly, the shoot tissues of transgenic
seedlings contained about 1.5- to 2.5-fold higher Cd(II) and Pb(II) levels than wild type (WT) seedlings, demonstrating significantly-increased
accumulation of both Cd(II) and Pb(II) in transgenic plants. The enhanced capacity of heavy metal tolerance and accumulation
by AtATM3 transgenic plants was attributed to higher BjGSHII (B. juncea glutathione synthetase II) and BjPCS1 (phytochelatin synthase 1) expression levels induced by AtATM3 overexpression. In addition, AtATM3 overexpression regulated the expression of several metal transporters in the transgenic B. juncea plants under heavy metal stress conditions. Therefore, AtATM3 transgenic plants are more tolerant of and can accumulate more heavy metals to enhance phytoremediation of contaminated soils.
Full-text · Article · Oct 2011 · Plant Cell Tissue and Organ Culture
[Show abstract][Hide abstract] ABSTRACT: The 5S rRNA gene unit was characterized from Lilium species distributed in Korea. Sequence analyses revealed that the 5S rDNA, harboring a highly conserved coding region (120
bp) and a divergent non transcribed spacer (NTS), ranged from 470 bp to 640 bp. The NTS regions showed length heterogeneity
(350 bp–520 bp) due to the presence of several indels, and extensive sequence divergence among the species. Despite heterogeneity
in length and nucleotide composition, the NTS regions possess some common features across the species, which include a T stretch
region CTTTT and an identical motif of 11 bp (CAATGTATGAC) at the 3′ and 5′ flanking sequence of the coding region, respectively.
These may play a role in the initiation and termination of transcription. The chromosomal distribution of the 5S rRNA gene
was mapped on the long arm of chromosome 3 by fluorescence in situ hybridization (FISH). The phylogenetic analysis based on the NTS sequence broadly divided the Lilium species into two major groups which were referred to as the section Sinomartagon and Martagon by Comber’s classification of the genus Lilium. The present study showed that the 5S rDNA sequence was very useful to unravel the genetic relationships among Lilium species.
–5S rRNA gene–NTS–genetic variation–FISH mapping
[Show abstract][Hide abstract] ABSTRACT: An efficient Agrobacterium-mediated genetic transformation method was established for Brassica juncea by investigating several factors responsible for successful gene transfer. Four-day-old cotyledon explants from in vitro grown seedlings were co-cultivated with Agrobacterium strain GV3101 harboring the binary vector EnPCAMBIA1302-YCF1, which contained the hygromycin phosphotransferase (HPT) gene as a selectable marker and the yeast cadmium factor 1 (YCF1) gene. Two days co-cultivation period on shoot induction medium (MS medium supplemented with 0.1 mg l(-1) alpha-naphthaleneacetic acid, 1.0 mg l(-1) 6-benzyladenine, and 2.0 mg l(-1) silver nitrate) containing 20 mg l(-1) acetosyringone and five days delaying exposure of explants to selective agent enhanced transformation efficiency significantly. A three-step selection strategy was developed to select hygromycin resistant shoots. Hygromycin-resistant shoots were subsequently rooted on root induction medium. Rooted plantlets were transferred to pot-soil, hardened, and grown in a greenhouse until maturity. Using the optimized transformation procedure, transformation efficiency reached at 16.2% in this study. Southern blot analysis was performed to confirm that transgenes (HPT and YCF1) were stably integrated into the plant genome. All transgenic plants showed single-copy of transgene integration in the host genome. Segregation analysis of T(1) progeny showed that the transgenes were stably integrated and transmitted to the progeny in a Mendelian fashion.
Full-text · Article · Jan 2011 · Plant Biotechnology
[Show abstract][Hide abstract] ABSTRACT: Molecular cytogenetic analyses using fluorescence in situ hybridization (FISH) and genomic in situ hybridization (GISH) were carried out to elucidate inter-specific relationships among wild Lilium species distributed in Korea. FISH revealed four to eight 45S rRNA gene loci, which are located on chromosomes 1–7, 10, and 11 among the different species. In contrast, the 5S rRNA gene locus was conserved on the long arm of chromosome 3, occasionally with two adjacent sites on the same chromosome arm in a few species. The 5S rDNA site was located adjacent to the 45S rDNA site in only three species, Lilium distichum, Lilium hansonii, and Lilium tsingtauense. GISH analysis using genomic DNA probes detected strong hybridization of genomes between diploid and triploid Lilium lancifolium species, demonstrating that triploid plants were derived from diploid L. lancifolium and not from Lilium maximowiczii. Phylogenetic analysis of the ITS and NTS sequences supported the cytogenetic data as well as Comber’s classification of the genus Lilium.
Preview · Article · Dec 2010 · Journal of Plant Biology
[Show abstract][Hide abstract] ABSTRACT: A yeast cadmium factor 1 (YCF1) is a member of the ATP-binding cassette (ABC) transporter family associated with multi-drug
resistance, and it is localized at the vacuolar membrane in Saccharomyces cerevisiae. To determine ability to increase heavy metal tolerance and accumulation, YCF1 was introduced into Brassica juncea plants by Agrobacterium-mediated genetic transformation. YCF1 gene presence in transgenic plants was demonstrated by polymerase chain reaction (PCR). Reverse transcriptase-PCR analysis
confirmed YCF1 gene expression in the transgenic plants, but the degree of YCF1 expression varied among the lines. YCF1 overexpression in B. juncea conferred enhanced tolerance to cadmium (Cd[II]) and lead (Pb[II]) stress. Transgenic B. juncea seedlings showed 1.3- to 1.6-fold tolerance to Cd stress and 1.2- to 1.4-fold tolerance to Pb stress compared to wild type
(WT) plants (per gram fresh weight). Most importantly, the shoot tissues of transgenic seedlings contained about 1.5- to 2-fold
higher Cd(II) and Pb(II) levels than those of WT, demonstrating significantly increased accumulation of both Cd(II) and Pb(II)
in transgenic plants.
–Phytoremediation–Heavy metal–ABC transporter–Yeast cadmium factor 1
Full-text · Article · Apr 2010 · Plant Cell Tissue and Organ Culture