1Department of Biology, Faculty of Sciences, Toyama University, Toyama 930-0887, 2Center for Gene Research, Nagoya University, Nagoya 464-8602, Japan
We have chosen whisk fern (Psilotum nudum) as the fifth target for our analysis of plastid genomes, because it is classified between Bryophyta and Gymnosperms. As it was practically impossible to isolate plastids from the hard plant or to separate the plastid DNA from a total DNA preparation, we amplified plastid DNA pieces by a long PCR technique. Eleven pairs of primers were designed based on the sequence of tobacco plastid DNA, and the plastid DNA fragments were amplified from the total DNA preparation of P. nudum cv. Kingyoku. Eleven PCR-fragments were found to cover the entire plastid genome. Each PCR-fragment was sonicated and size- fractionated. The resultant DNA pieces (average 1.5 kbp) were cloned into Bluescript KS+ and sequenced. Sequence data were assembled into a circle and the total length of the genome was found to be 138,829 bp. P. nudum plastid DNA has a large inverted repeat of 18,954 bp and shows an intermediate feature in gene organization between Bryophyta and Gymnosperm (1). The overall gene organization is similar to that of tobacco, except the 4.5 kbp region from "trnT -- psbDC -- trnG" is inversely oriented. It contains four different rRNA genes and 32 different tRNA genes including Pro-tRNA (GGG) gene (this was found only in black pine, 2). Seventy three different protein-coding genes and 8 ycfs (conserved ORFs) have so far been identified. The plastid genome has a gene encoding ribosomal protein L21, which was found as a plastid gene only in Marchantia plastids. The ribosomal protein S12 genes are divided genes in most land plant plastids and contain one trans-intron and one cis-intron (1). The corresponding gene in P. nudum plastids is also a divided gene but harbors no cis-intron, which is a feature unique to the plastid genome from P. nudum and Cuscuta europaea (a parasitic land plant, 3).
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