Use of the polymerase chain reaction to detect spacer size heterogeneity in plant 5S-rRNA gene clusters and to locate such clusters in wheat (Triticum aestivum L.)

• Published in: Theoretical and applied genetics 1992-04, Vol.83 (6-7), p.684-690
• Main Authors: Cox, A.V. (Royal Botanic Gardens, Kew (United Kingdom). Jodrell Lab.), Bennett, M.D, Dyer, T.A
• Format: Article
• Language: English
• Subjects: 5S-ribosomal RNA; ARN RIBOSOMAL; ARN RIBOSOMIAL; Biological and medical sciences; Chromosome location; Fundamental and applied biological sciences. Psychology; GENE LOCATION; Genes. Genome; GENETIC POLYMORPHISM; LOCALISATION DE GENE; LOCALIZACION DE GENES; Molecular and cellular biology; Molecular genetics; Non-transcribed spacer; NUCLEOTIDE SEQUENCE; PCR; POLIMORFISMO GENETICO; POLYMORPHISME GENETIQUE; RIBOSOMAL RNA; SECUENCIA NUCLEICA; SEQUENCE NUCLEIQUE; TRITICUM AESTIVUM
• ISSN: 0040-5752; 1432-2242
• DOI: 10.1007/BF00226685

We have used the polymerase chain reaction to analyse variation in the size of individual 5S-ribosomal gene spacer sequences. This reaction can be used to demonstrate inter- and intraspecific variation in spacer size, and combined with DNA sequencing it may thus be a valuable taxonomic tool. Two sets of nested polymerase chain reaction primers were designed to amplify the nontranscribed spacer DNA between repeated 5S-rRNA genes. These "universal" primers were used to generate fragments from the genomic DNA from several unrelated monocotyledonous plants. Ribosomal RNA spacer sequences generated in these experiments could also be used to locate 5S-rRNA gene clusters on specific chromosomes in hexaploid wheat (Triticum aestivum). Three distinct spacer sizes were observed after amplification. These were assigned locations on chromosomes by analysing amplification products of genomic DNA from nullisomic/tetrasomic and ditelosomic wheat stocks. "Large" 508-bp 5S repeats are located on the short arm of chromosome 5B and "short" 416-bp and 425-bp repeat unit variants are located on the short arms of chromosomes 1B and 1D, respectively. No other loci were detected. The spacer fragments were cloned, sequenced, and shown to be homologous to wheat 5S-rRNA spacers previously identified. Spacers of uniform size but with some sequence heterogeneity were shown to be located at each locus.