Cloning And Sequencing Of Several Segments Of Rice Gall Dwarf Virus Genome

The nucleotide sequences of DNAs complementary to the part segments of the 12 genome segments of rice gall dwarf virus Xinyi isolate (RGDV-C) have beencloned and their sequences have been determined. The fragments corresponding tothe terminal and conserved sequences of rice gall dwarf virus Thailand isolate (RGDV-T) were amplified by RT-PCR from RGDV-C genome and sequenced. The complete nucleotide sequences of each genome segment were assembled and deposited in the GenBank databases. The accession numbers are AY556484 (S2) , AY559408 (S3), AY216767 (S8), AY556483 (S9), AY559406 (S10), AY559407 (S11) , respectively. Genome segment between RGDV-C and RGDV-T shared 91.9%-98.1% and 96.6%-98.7% homology at level of nucleotide and amino acid sequence, respectively. From multiple nucleotide sequence alignments, it was found that most site mutations were base transitions (A/G or C/T substitution) and most mutation occurred on the third base of codons in the open reading frames (ORFs). At the level of amino acid sequences, most mutations were substitution between the similar amino acids. It was noticeable that only one deletion of Leu occurred at position aa 211 in the ORF of RGDV-C S11, which suggestion the deletion did not affect the function of the protein. The genomic sizes and features of RGDV-C were very similar to those of corresponding segments of RGDV-T. From the Phylogenetic relationships of outer coat protein and the virus particles are restricted to the phloem and gall cells of infected plants, RGDV are more similar to those of WTV than those of RDV.The nucleotide sequences of S12 have been determined by the method of single primer amplification firstly. The full-length cDNA of S12 has 853 nucleotides, and shows a common characteristic feature of terminal sequence (5'GG-UGAU 3'). Some inverted repeat sequence appears in RGDV-C genome segment S12. Alignment of the complete nucleotide sequences of S12 among RGDV, RDV and WTV, they shared approximately 50% homologous. The computer analysis demonstrate that RGDV S12 mRNA maybe contain two open reading frame andspecified translation products from the large ORF and one overlapping small ORF. These provide the first example of a bicistronic mRN A for a segment hi ROD V.Computer-assisted search analyses have detected conserved domains in this family consisted of several Phytoreovirus outer capsid protein P8 sequences; The family consisted of the Phytoreovirus nonstructural proteins Pns9 and PnslO have conserved domains too, but the function of this family is unknown. Phytoreovirus nonstructural protein PnslO/11 family shared the similar conserved architecture with Phage integrase family, whose member cleave DNA substrates by a series of staggered cuts, during which the protein becomes covalently linked to the DNA through a catalytic tyrosine residue at the carboxy end of the alignment. Pns9 protein encoded by RGDV S9 shared 21.8% homology with ATP-dependent Clp protease proteolytic component at level of ammo acid sequence.