1988. Apparent sources of the A genomes of wheats inferred from polymorphism in abundance and restriction fragment length of repeated nucleotide sequences. Genome, 30: 680-689. Four hundred random DNA fragment clones of wild diploid wheat Triticum monococcum ssp. aegilopoides (syn. T baeoticum) were screened for clones of repeated nucleotide sequences. Seven DNA fragments were isolated that were more abundant by one order of magnitude or more in the genome of diploid T monococcum ssp. aegilopoides (genome A) than in the genome of diploid Triticum speltoides (genome BS). These clones were then used to determine which of the two wild diploid wheats, T m. ssp. aegilopoides or T urartu, was the ancestor of domesticated diploid wheat T m. ssp. monococcum, wild tetraploid wheats T turgidum ssp. dicoccoides and T timopheevii ssp. araraticum, domesticated tetraploid wheat T turgidum, and hexaploid bread wheat T aestivum. Three of the seven cloned repeated nucleotide sequences differentiated the genome of T m. ssp. aegilopoides from that of T urartu in repeated sequence abundance, restriction fragment length polymorphism, or both. The same distinctions were observed between the A genome of T m. ssp. aegilopoides and the A genomes of polyploid wheats. From this it was concluded that the species from which T m. ssp. monococcum was domesticated was T m. ssp. aegilopoides but that the A genomes of the polyploid wheats are equivalent to that of T urartu. The results presented here demonstrate the utility of polymorphism in repeated nucleotide sequences in the investigation of the origin of genomes in polyploid plants.Key words: RFLF, Triticum, wheat phylogeny.DVORAK, J., MCGUIRE, P. E., et CASSIDY, B. 1988. Apparent sources of the A genomes of wheats inferred from polymorphism in abundance and restriction fragment length of repeated nucleotide sequences. Genome, 30 : 680-689. Quatre cents clones de fragments d'ADN, pris au hasard, du blC diplo'ide sauvage Triticum monococcum ssp. aegilopoides (syn. T baeoticum) ont Ct C examines pour dCceler les clones possCdant des sequences rCpCtCes de nuclCotides. Sept fragments d'ADN ont Ct C isolCs, qui representaient de fa~on plus abondante le gCnome T m. aegilopoides (genome A) plut6t que le genome T speltoides (gCnome BS), par au moins un ordre de grandeur ou davantage. Ces clones furent utilisCs pour determiner lequel des deux bles sauvages, T m. ssp. aegilopoides ou T urartu, Ctait l'ancstre du blC diplo'ide T m. ssp. monococcum introduit en culture, des blCs tktraplo'ides sauvages T turgidum ssp. dicoccoides et T timopheevii ssp. araraticum, du blC tktraplo'ide cultivC T turgidum et du blC panifiable hexaplo'ide T aestivum. Trois des sept sequences de nuclCotides rCpCtCes et clonees ont differencie le gCnome T m. ssp. aegilopoides du genome T urartu en fonction de l'abondance des sequences rCpCtCes, du polymorphisme de la longueur des fragments de restriction (PLFR = RFLP), ou des deux. Les msmes distinctions ont Ct C observCes entre le gCnome A de T m. ssp. aegilopoides et les gCno...

No abstract

The DNA sequence of the intragenic region of the rat 45S ribosomal RNA precursor was determined. This sequence contains 2282 nucleotides and extends from the conserved EcoR I site near the 3' terminus of 18S rRNA to 69 nucleotides downstream of the 5' terminus of 28S rRNA. The sequences corresponding to 18S and 5.8S rRNA were identified by comparison with previously published data. The 5' terminus of rat 28S rRNA was identified by S1 nuclease protection and reverse transcriptase elongation assays. The internal transcribed spacers were found to be 1066 and 765 nucleotides long and had little homology with those of Xenopus and yeast. Regions of sequence homology between rat and Xenopus were found at the junctions of the internal transcribed spacers with 18S, 5.8S and 28S rRNA. These homologies suggest that these sequences may function as recognition sites for the processing of the ribosomal precursor RNA.

In vitro transcription of the rat rRNA gene led to the identification of a region within a 3.4-kilobase fragment of the nontranscribed spacer (NTS) which significantly increased the transcription of rat ribosomal DNA. Promoter constructs containing this region were transcribed up to 17-fold more efficiently in vitro than templates with only 167 or 286 base pairs of NTS. This effect was also observed when the 3.4-kb fragment of the NTS was subcloned in the opposite orientation and 4 kb upstream of the promoter. The region responsible for the enhanced level of transcription was found between -286 and -1018. The results of order-of-addition experiments suggested that the enhanced level of transcription was the result of the formation of a stable complex between a trans-acting factor and the nontranscribed spacer. DNA-protein binding assays demonstrated that the same region of the NTS determined to have enhancer activity also specifically bound a proteinase K-sensitive factor present in nuclear extracts. The sequence of this region was not found to have any significant homology with the promoter of the rat rRNA gene. This is the first report to assign a transcriptional role to the NTS of a mammalian rRNA gene.

We identified and characterized an additional promoter within the nontranscribed spacer (NTS) of the rat ribosomal gene repeat that is capable of supporting initiation of transcription by RNA polymerase I in vitro. Within this promoter there is a sequence of 13 nucleotides which is 100% homologous to nucleotides -18 to -6 (+1 being the frst nucleotide of 45S rRNA) of the major promoter of 45S pre-rRNA and is located between nucleotides -731 and -719. To identify the exact location of the upstream initiation site, the RNA synthesized in vitro from this new promoter was gel isolated and subjected to fingerprint analysis, Southern hybridization, and reverse transcriptase elongation. Based on these analyses, the in vitro-synthesized RNA initiates with an A at nucleotide -713. When compared individually, the upstream promoter was transcribed ninefold less efficiently than the major promoter. When templates which contain both promoters on the same piece of DNA were transcribed, the major promoter was at least 50-fold more efficient.Eucaryotic ribosomal genes are arranged as tandemly repeating units. The transcription units that code for the RNA precursor are flanked by the nontranscribed spacer (NTS) regions. The transcribed portion of the repeat consists of an external transcribed spacer, the region that codes for 18S rRNA, an internal transcribed spacer, the 5.8S coding region, a second internal transcribed spacer, the region that codes for 28S rRNA, and a short 3' external transcribed spacer (23).Besides functioning to direct and regulate transcription, the NTS has been both proposed and shown to have roles in recombination (39), DNA replication (37), and chromatin structure (8).By far the greatest number of studies on the NTS have focused on its role in transcription. The DNA sequences immediately upstream of the rRNA transcription initiation site have been intensively studied by several laboratories. A core promoter region, located between nucleotides -39 and +6, has been identified, and specific nucleotides within this region have been shown to be required for initiation (11, 18, 2,, 25, 38, 43, 47). A second region, the upstream control element or upstream promoter element (between nucleotides 150 and -110), has been demonstrated to stabilize effectively the preinitiation complex (15,19,30,44,45; B. G. Cassidy, R. Haglund, and L. I. Rothblum, Biochim. Biophys. Acta, in press).In yeasts and Xenopus laevis, regions several hundred to several thousand nucleotides upstream of the 5' end of the ribosomal precursor have been shown to have profound effects on promoter utilization and the efficiency of transcription. In yeasts, a polymerase I promoter (42) or enhancer element (9) has an effect on the transcription of the 35S rRNA precursor (10). Two promoter-related elements present in multiple copies in the NTS of X. laevis have also been identified (reviewed in references 7 and 34). These elements, the Bam islands and the 60-or 81-base-pair (60/81-bp) repeats, affect the utilization of the polymerase I promoter and ...

Aphid transmission of potyviruses depends on the presence of specific sequence domains in two virus encoded proteins, the coat protein (CP) and helper component-proteinase (HC-Pro). Aphid transmissable peanut stripe virus (PStV), like most potyviruses, has an Asp-Ala-Gly (DAG) motif in the amino-terminal part of the CP. Peanut Mottle Virus (PeMoV) was determined to be highly aphid transmissible but has a unique Asp-Ala-Ala-Ala (DAAA) motif. To determine if the DAAA motif could functionally replace the DAG motif in PStV, mutations were made in a full-length cDNA clone of PStV. All of the mutations in the CP DAG motif abolished aphid transmissibility of PStV but did not affect virus infectivity. The aphid transmissibility of the PStV-DAAA mutant was partially restored by feeding aphids an artificial diet containing purified virus and PeMoV HC-Pro. The PStV-DAAA virus was poorly transmitted by aphids in vitro with HC-Pro purified from PStV or tobacco vein mottling virus (TVMV) infected plants. These experiments support the theory that specific HC-Pro/CP interactions are required for efficient aphid transmission. Based upon the sequence comparisons of 16 potyviral HC-Pro proteins several conserved motifs and striking differences have been identified. PeMoV was determined to have an Ala-Ser-Cys (ASC) HC-Pro motif instead of a highly conserved Cys-Cys-Cys (CCC) motif. We have predicted that this CCC motif could play an important role in the specific interaction between the HC-Pro and the CP DAG motif.

No abstract

A full-length, low-molecular-weight (LMW) glutenin cDNA clone, pTdUCD1, has been isolated from a Triticum durum cv 'Mexicali' wheat cDNA library. The complete sequence was determined and compared to the LMW glutenin genes that have been isolated from hexaploid wheat, Triticum aestivum. This cDNA codes for a protein of 295 amino acids (33,414 daltons) including a 20-amino acid signal peptide as deduced from the DNA sequence. Northern analysis showed that this cDNA hybridizes to a family of related sequences ranging in length from 1,200 to 1,000 nucleotides. This gene is similar but not identical to previously published LMW glutenin gene sequences. The most striking characteristic of all cloned LMW glutenin genes is the conservation of eight cysteine residues, which could be involved in potential secondary or tertiary structure, disulfide bond interactions. This paper presents a structural map defining distinct regions of the LMW glutenin gene family.