Plastid transformation reveals that moss tRNAArg-CCG is not essential for plastid function

Three distinct arginine tRNA genes, trnR-CCG, trnR-ACG, and trnR-UCU, are present in the plastid genome of bryophytes, whereas only the latter two trnR genes are present in the major vascular plants, except for black pine. trnR-CCG is located between rbcL and accD in the moss Physcomitrella patens and it was previously believed to be functional in plastids. However, no trnR-CCG transcript has been detected by Northern hybridization, and the codon usage of CGG is quite low in plastid protein-coding sequences. This raises the possibility that trnR-CCG is non-functional. To investigate this possibility, we integrated a foreign gene into the trnR-CCG coding region via homologous recombination, and constructed stable plastid trnR-CCG knock-out moss transformants. The trnR-CCG knock-out transformants grew normally, indicating that the P. patenstrnR-CCG gene is not essential for plastid function.