Hairong Wei

Rapid detection of Prunus necrotic ringspot virus using magnetic nanoparticle-assisted reverse transcription loop-mediated isothermal amplification.

Prunus necrotic ringspot virus (PNRSV) has seriously reduced the yield of Prunus species worldwide. In this study, a highly efficient and specific two-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) was developed to detect PNRSV. Total RNA was extracted from sweet cherry leaf samples using a commercial kit based on a magnetic nanoparticle technique. Transcripts were used as the templates for the assay. The results of this assay can be detected using agarose gel electrophoresis or by assessing in-tube fluorescence after adding SYBR Green I. The assay is highly specific for PNRSV, and it is more sensitive than reverse-transcription polymerase chain reaction (RT-PCR). Restriction enzyme digestion verified further the reliability of this RT-LAMP assay. To our knowledge, this is the first report of the application of RT-LAMP to PNRSV detection in Prunus species.

Complete nucleotide sequence of little cherry virus 1 (LChV-1) infecting sweet cherry in China

Little cherry virus 1 (LChV-1), associated with little cherry disease (LCD), has a significant impact on fruit quality of infected sweet cherry trees. We report the full genome sequence of an isolate of LChV-1 from Taian, China (LChV-1-TA), detected by small-RNA deep sequencing and amplified by overlapping RT-PCR. The LChV-1-TA genome was 16,932xa0nt in length and contained nine open reading frames (ORFs), with sequence identity at the overall genome level of 76xa0%, 76xa0%, and 78xa0% to LChV-1 isolates Y10237 (UW2 isolate), EU715989 (ITMAR isolate) and JX669615 (V2356 isolate), respectively. Based on the phylogenetic analysis of HSP70h amino acid sequences of Closteroviridae family members, LChV-1-TA was grouped into a well-supported cluster with the members of the genus Velarivirus and was also closely related to other LChV-1 isolates. This is the first report of the complete nucleotide sequence of LChV-1 infecting sweet cherry in China.

Transcriptome Analysis, Marker Discovery and Pigment Biosynthesis of Red-leaf Juglans regia

Red-leaf trait rarely occurs in Juglans regia , and the genetic mechanism underlying this phenomenon is still unknown. In this study, we attempted to provide insight into the comprehensive transcriptome of red-leaf J. regia by RNA-Seq using Illumina SeqTM2000 platform. A total of 33,488,602 high-quality reads (3.35G cleans bases) were obtained and assembled into 53,782 unigenes. A total of 3,683 unigenes were annotated by using basic local alignment search tool to search against protein databases, All the matched unigenes were categorized by gene ontology analysis, and 3,466 were assigned to metabolism, among which 74 were mapped to anthocyanin, carotenoid, and betalain biosynthetic pathways by Kyoto Encyclopedia of Genes and Genomes analysis. Approximately 656 transcription factors were isolated including MYB, NAC, and bHLH. Additionally, a total of 13,981 simple sequence repeats, 41,088 single nucleotide polymorphisms, and 5,860 insertions and deletions were determined from J. regia transcriptome. Therefore, the current J. regia transcriptome provides deep insight into the molecular basis of red-leaf breeding of J. regia .

Characterising novel S-RNase and SFB alleles in Prunus virginiana

Summary Self-incompatibility (SI) in the genus Prunus is controlled by alleles of two genes at the S-locus, S-RNase and SFB. Both genes exhibit high levels of polymorphism and sequence diversity, characteristic of plant self-incompatibility systems. In this study, eight S-RNase alleles and five SFB alleles were isolated from five accessions of P. virginiana using PCR. These alleles were novel and have been submitted to GeneBank with the following Accession Numbers: S1-RNase – S8-RNase (JF907557 – JF907564), SFB1 (JF907565), SFB2 (JF907566), and SFB4 (JF907567). The Accession Numbers of SFB7 and SFB8 are the same as S7-RNase and S8-RNase, respectively. These S-RNase and SFB alleles shared the typical structural features of the equivalent alleles in other Prunus species. The identities of the deduced amino acid sequences range from 70.5 – 80.9% among the S-RNase alleles, and from 66.5 – 82.9% among the SFB alleles. The S-RNase and SFB genes were transcribed in opposite directions. The distances between pairs of alleles in these S-haplotypes varied from 325 – 1,789 bp. Sequence comparisons showed that the identities of the deduced amino acid sequences were 99.4% between Pvir-S3 and Pweb-S10, 99.0% between Pvir-S3 and Pd-S64, 97.1% between Pvir-S7 and Pps-S5, 98.2% between Pvir-S4 and Pa-S6, 98.2% between Pvir-S6 and Pspe-S3, 98.4% between Pvir-SFB4 and Pa-SFB6, and 97.4% between Pvir-SFB7 and Pps-SFB5. Moreover, the identity between the intergenic sequences of the S7-haplotype of P. virginian and the S5-haplotype of P. pseudocerasus was 99.7%, with only one nucleotide difference (C → T). These results show that these two S-haplotypes were the closest related lineages and further support the view that alleles at these two S-loci evolved before the divergence of Prunus species.