ITS1 sequences of nuclear ribosome DNA in rice species from China and their phylogenetic implications

Abstract

This study assessed the capability of using internal transcribed spacer (ITS1) sequences for reconstructing the phylogeny of rice species from China. Three accessions of wild and two accessions of cultivated rice were used (Table 1). Total DNA was extracted from leaf samples following the procedures described by Tai and Tanksley (1990). Polymerase chain reaction (PCR) amplifications were performed in 50 μL volumes containing 100 ng DNA template and 2.5 μL Taq polymerase. A PE-480 thermal cycler was used under the following conditions: 35 cycles of 94°C for 1 min, 57°C for 1 min, and 72°C for 1 min. PCR-amplified DNA products were recovered, complemented, and then cloned into the vector pUC 118 with standard protocols. Sequencing of ITS1 was conducted on an AB1 373A automated sequencer using a TaqDye Primer Cycle Sequencing Kit (ABI). The ITS1 sequences of the five taxa together with a published rice sequence (Takaiwa et al 1985) were aligned with the Clustal V program. The data matrix was analyzed using UPGMA with Jukes-Cantor distance and Neighbor-Joining with Kimura 2-parameter distance of the MEGA version 1.02 (Kura et al 1993).