Characterization and Expression Analysis of Four Glycine-Rich RNA-Binding Proteins Involved in Osmotic Response in Tobacco (Nicotiana tabacum cv. Xanthi)

Plants have developed many signals and specific genes＇ regulations at both transcriptional and post-transcriptional levels in order to tolerate and adapt to various environmental stresses. RNA-binding proteins （RBPs） play crucial roles in the post- transcriptional regulation via mRNA splicing, polyadenylation, sequence editing, transport, mRNA stability, mRNA localization, and translation. In this paper, four cDNAs of glycine-rich RNA-binding proteins （GR-RBPs）, named NtRGP-la, -lb, -2, and -3, were isolated from Nicotiana tabacum by RT-PCR analysis, and special emphases were given to the sequences alignment, phylogenetic analysis and gene expression. Sequences alignment revealed minor difference of cDNA sequences, but no difference of deduced proteins between N. sylvestris and N. tabacum. Phylogenetic alignment revealed that four cDNAs in tobacco were clustered into two different groups. NtRGP-2 and -3 were evolutionarily closest to Arabidopsis GR-RBPs genes and related to animal GR-RBPs genes, while NtRGP-la and -lb were closest to Gramineae GR-RBPs genes. The expression analyses of these four NtRGPs in response to different abiotic stresses revealed the similar expression pattern. Moreover, the four NtRGPs, especially NtRGP-la and NtRGP-3, were strongly induced by stresses including water, wound, cold, and high temperature, weakly induced by PEG, drought and SA, while reduced by NaC1 and unaffected by ABA treatment. The fact that all of these abiotic stresses included in our experiments affected the water balance and resulted in osmotic stress on cellular level, suggests that NtRGPs in tobacco should be a family of crucial osmosis-related proteins, and may play a key role in signal transduction with ABA-independent pathway under abiotic stresses.

Nucleotide Sequencing and Phylogenetic Analysis Using PCR Amplicons of U3 Gene of Jaagsiekte Sheep Retrovirus (JSRV) Detected in Natural Cases of Ovine Pulmonary Adenocarcinoma in India

Ovine pulmonary adenocarcinoma (OPA) caused by an exogenous Jaagsiekte sheep retrovirus (JSRV) is prevalent in Indian sheep. In the present study, OPA was diagnosed in sheep by clinical signs, gross and histopathology and polymerase chain reaction (PCR). Proviral DNA of exogenous JSRV was detected in lung tumor tissues, mediastinal lymph nodes, blood and lung fluid samples from natural cases of OPA by using U3-hn PCR and the PCR amplicons were sequenced to analyze nucleotide divergence. In total, six isolates were sequenced that had 96% - 100% homology with a UK strain (AF105220.1) but more divergent from a South African strain (M80216) with 88% - 93% identity. The phylogenetic analysis revealed segregation of the six isolates into two clusters. In conclusion, this study is the first report on sequencing and phylogenetic analysis of JSRV in India and further studies are suggested to know the complete sequencing and genetic divergence of JSRV in Indian sheep.

Identification and Assessing the Cultivars of Laminaria Lamx. （Phaeophyceae） with Molecular Markers

Molecular markers were used to identify and assess cultivars of Laminaria Lamx. and to delineate their phylogenetic relationships. Random amplified polymorphic DNA (RAPD) analysis was used for detection. After screening, 11 primers were selected and they yielded 133 bands in all, of which approximately 99.2% were polymorphic. The genetic distances between gametophytes ranged from 0.412 to 0.956. Two clusters were formed with the unweighted pair group method with arithmetic mean (UPGMA) dendrogram based on the simple matching coefficient. All cultivars of Laminaria japonica Aresch. used for breeding in China fell into one cluster. L. japonica from Japan, L. saccharina (L.) Lam., and L. angustata Kjellm. formed the other cluster and showed higher genetic variation than L. japonica from China. Nuclear ribosomal DNA (rDNA) sequences, including internal transcribed spacers (ITS 1 and ITS2) were studied and aligned. The nucleotides of the sequences ranged from 634 to 668, with a total of 692 positions including ITS1, ITS2, and the 5.8S coding region. The phylogenetic tree obtained by the neighbor-joining method favored, to some extent, the results revealed by RAPD analysis. The present study indicates that RAPD and ITS analyses could be used to identify and assess Laminaria germplasm and to distinguish some species and, even intraspecies, in Laminaria.

Analysis of Diversity of CO I Gene and System Generation of Nine Species of Chitons

Cytochrome c oxidase subunit 1 （CO I ） genes from 9 chiton species in China＇s coast area were sequenced. A phylogeny tree about these gene sequences were reconstructed together with other 4 CO I gene sequences of chitons from Genbank. The affin- ity genetic relationship and the taxonomic status of molecular evolution for these 13 chiton species were analyzed. The results show that Liolophura japonica and Onithochiton hirasei belong to Chitonidae, .4canthochiton rubrolineatus and Acanthochiton dis- similis belong to Acanthochitonidae, and Placiphorella japonica and Mopalia retifera belong to the same family Mopliidae. How- ever, Lepidozona coreanica, Ischnochiton comptus, and lschno- chiton hakodadensis are not supposed to be referred to Ischnochi- tonidae according to the genetic distance analysis （L. coreanica, 1. comptus, and 1. hakodadensis are usually classed as Ischnochi- tonidae according to their morphological character）. Furthermore, 1. hakodadensis could not been classed as lschnochiton, and it is more likely to be treated as a close relative of Lepidozona.

SARS-CoV Genome Polymorphism: A Bioinformatics Study

A dataset of 103 SARS-CoV isolates （101 human patients and 2 palm civets） was investigated on different aspects of genome polymorphism and isolate classification. The number and the distribution of single nucleotide variations （SNVs） and insertions and deletions, with respect to a “profile”, were determined and discussed （“profile” being a sequence containing the most represented letter per position）. Distribution of substitution categories per codon positions, as well as synonymous and non-synonymous substitutions in coding regions of annotated isolates, was determined, along with amino acid （a.a.） property changes. Similar analysis was performed for the spike （S） protein in all the isolates （55 of them being predicted for the first time）. The ratio Ka/Ks confirmed that the S gene was subjected to the Darwinian selection during virus transmission from animals to humans. Isolates from the dataset were classified according to genome polymorphism and genotypes. Genome polymorphism yields to two groups, one with a small number of SNVs and another with a large number of SNVs, with up to four subgroups with respect to insertions and deletions. We identified three basic nine-locus genotypes： TTTT/TTCGG, CGCC/TTCAT, and TGCC/TTCGT, with four subgenotypes. Both classifications proposed are in accordance with the new insights into possible epidemiological spread, both in space and time.

Cloning and Preliminary Characterization of Three Receptor-like Kinase Genes in Soybean

Leaf senescence that occurs in the last stage of leaf development is a genetically programmed process. It is very significant to isolate the upstream components in the senescence signaling pathway and to elucidate the molecular mechanisms that control the initiation and progression of leaf senescence. In this study, full-length cDNAs of three receptor-like protein kinase genes, designated rlpkl, rlpk2 and rlpk3, were cloned from artificially-induced senescent soybean （Glycine max L.） primary leaves （GenBank accession AY687390, AY687391, AF338813）. The deduced amino acid sequences indicated that they belonged to a receptor-like kinase family. Each of rlpkl and rlpk2 encodes a leucine-rich repeat （LRR） receptor-like protein kinase. They both comprise a typical signal peptide, several LRR motifs, a single-pass transmembrane domain, and a cytoplasmic protein kinase domain. No typical extracellular domain of RLPK3 was predicted. Organ-specific expression pattern analysis by reverse-transcription polymerase chain reaction （RT-PCR） revealed higher expression levels of the three genes in cotyledons, roots and flowers. Phylogenetic analysis indicated that RLPK1 and RLPK2 belonged to an independent branch, whereas RLPK3 shared common nodes with several known RLKs responding to ablotic and biotic stresses. The evident alternations of expression profiles of rlpkl and rlpk2 induced by the artificial senescence-inducing treatment implied involvements of these two RLKs in regulating soybean leaf senescence.