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Complete chloroplast genome of Oncidium Gower Ramsey and evaluation of molecular markers for identification and breeding in Oncidiinae.

ABSTRACT: BACKGROUND:Oncidium spp. produce commercially important orchid cut flowers. However, they are amenable to intergeneric and inter-specific crossing making phylogenetic identification very difficult. Molecular markers derived from the chloroplast genome can provide useful tools for phylogenetic resolution. RESULTS:The complete chloroplast genome of the economically important Oncidium variety Onc. Gower Ramsey (Accession no. GQ324949) was determined using a polymerase chain reaction (PCR) and Sanger based ABI sequencing. The length of the Oncidium chloroplast genome is 146,484 bp. Genome structure, gene order and orientation are similar to Phalaenopsis, but differ from typical Poaceae, other monocots for which there are several published chloroplast (cp) genome. The Onc. Gower Ramsey chloroplast-encoded NADH dehydrogenase (ndh) genes, except ndhE, lack apparent functions. Deletion and other types of mutations were also found in the ndh genes of 15 other economically important Oncidiinae varieties, except ndhE in some species. The positions of some species in the evolution and taxonomy of Oncidiinae are difficult to identify. To identify the relationships between the 15 Oncidiinae hybrids, eight regions of the Onc. Gower Ramsey chloroplast genome were amplified by PCR for phylogenetic analysis. A total of 7042 bp derived from the eight regions could identify the relationships at the species level, which were supported by high bootstrap values. One particular 1846 bp region, derived from two PCR products (trnHGUG -psbA and trnFGAA-ndhJ) was adequate for correct phylogenetic placement of 13 of the 15 varieties (with the exception of Degarmoara Flying High and Odontoglossum Violetta von Holm). Thus the chloroplast genome provides a useful molecular marker for species identifications. CONCLUSION:In this report, we used Phalaenopsis. aphrodite as a prototype for primer design to complete the Onc. Gower Ramsey genome sequence. Gene annotation showed that most of the ndh genes inOncidiinae, with the exception of ndhE, are non-functional. This phenomenon was observed in all of the Oncidiinae species tested. The genes and chloroplast DNA regions that would be the most useful for phylogenetic analysis were determined to be the trnHGUG-psbA and the trnFGAA-ndhJ regions. We conclude that complete chloroplast genome information is useful for plant phylogenetic and evolutionary studies in Oncidium with applications for breeding and variety identification.

SUBMITTER: Wu FH

PROVIDER: S-EPMC3095342 | biostudies-literature | 2010 Apr

REPOSITORIES: biostudies-literature

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Complete chloroplast genome of Oncidium Gower Ramsey and evaluation of molecular markers for identification and breeding in Oncidiinae.

Wu Fu-Hui FH Chan Ming-Tsair MT Liao De-Chih DC Hsu Chen-Tran CT Lee Yi-Wei YW Daniell Henry H Duvall Melvin R MR Lin Choun-Sea CS

BMC plant biology 20100416

<h4>Background</h4>Oncidium spp. produce commercially important orchid cut flowers. However, they are amenable to intergeneric and inter-specific crossing making phylogenetic identification very difficult. Molecular markers derived from the chloroplast genome can provide useful tools for phylogenetic resolution.<h4>Results</h4>The complete chloroplast genome of the economically important Oncidium variety Onc. Gower Ramsey (Accession no. GQ324949) was determined using a polymerase chain reaction ...[more]

PMID: 20398375

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Project description:Orchids are commercially important flowers, comprising one of the largest families of plants. Model plants such as Arabidopsis do not contain all plant genes. In this study, several molecular biology tools were integrated to isolate the flower-specific gene promoters of Oncidium Gower Ramsey (GR). A cDNA library of reproductive tissues was constructed and a microarray was established and used to compare gene expression in flowers and leaves. Five genes were highly expressed in flower tissues and the subcellular locations of the corresponding proteins were identified using fluorescent protein-fusion protein lip transient transformation, and related to their putative functions. BAC clones of the five genes, together with nine previously published flower and reproductive growth specific genes in Onc. GR, were identified and promoter regions were cloned by screening an Onc. GR BAC library. Interestingly, three of the five novel flower-abundant genes were putative trypsin inhibitor (TI) genes, OnTI1, OnTI2 and OnTI3, which were tandemly duplicated in the same BAC clone. Their promoters were identified using transient GUS reporter gene transformation and stable Arabidopsis transformation analyses. In conclusion, a combination of tools including a cDNA microarray, a BAC library, and a bombardment assay can be successfully employed to identify orchid genes and promoters. A total of 960 cDNAs derived from the Onc. GR flower cDNA library were amplified using PCR incorporating the T3 and M13 reverse universal primers. The PCR products were purified using the QIAquick PCR Purification kit (Qiagen, Germantown, MD) and eluted into 100 M-BM-5l of 0.1M-CM-^W Tris-EDTA buffer, pH 8.0. Purified PCR products were printed on GAPS II-coated slides (Corning, New York, NY) using the OmniGrid 100 microarray (Genomic Solutions, Ann Arbor, MI), and arranged into 1.8 M-CM-^W 1.8-cm arrays (spot size: 100 M-NM-<m) twice. These DNAs were cross-linked to the slide by baking the array at 80M-BM-0C for 2 h. After printing, the slides were left to dry overnight. The total RNA from leaves and flowers (25 M-NM-<g) was used for cDNA synthesis with either Cy3 or Cy5 dye molecules, using the 3DNA Expression Array Detection kit for microarrays (Array 50, version 2, Genisphere, Hatfield, PA). cDNA hybridization and washing procedures were performed according to the manufacturer's instructions. All experiments were carried out in triplicate, with repeats of each dye combination (n = 3).

Project description:Among the four species of Echinacanthus (Acanthaceae), one distributed in the West Himalayan region and three restricted to the Sino-Vietnamese karst region. Because of its ecological significance, molecular markers are necessary for proper assessment of its genetic diversity and phylogenetic relationships. Herein, the complete chloroplast genomes of four Echinacanthus species were determined for the first time. The results indicated that all the chloroplast genomes were mapped as a circular structure and each genomes included 113 unique genes, of which 80 were protein-coding, 29 were tRNAs, and 4 were rRNAs. However, the four cp genomes ranged from 151,333 to 152,672 bp in length. Comparison of the four cp genomes showed that the divergence level was greater between geographic groups. We also analyzed IR expansion or contraction in the four cp genomes and the fifth type of the large single copy/inverted repeat region in Lamiales was suggested. Furthermore, based on the analyses of comparison and nucleotide variability, six most divergent sequences (rrn16, ycf1, ndhA, rps16-trnQ-UUG, trnS-GCU-trnG-UCC, and psaA-ycf3) were identified. A total of 37-45 simple sequence repeats were discovered in the four species and 22 SSRs were identified as candidate effective molecular markers for detecting interspecies polymorphisms. These SSRs and hotspot regions could be used as potential molecular markers for future study. Phylogenetic analysis based on Bayesian and parsimony methods did not support the monophyly of Echinacanthus. The phylogenetic relationships among the four species were clearly resolved and the results supported the recognition of the Sino-Vietnamese Echinacanthus species as a new genus. Based on the protein sequence evolution analysis, 12 genes (rpl14, rpl16, rps4, rps15, rps18, rps19, psbK, psbN, ndhC, ndhJ, rpoB, and infA) were detected under positive selection in branch of Sino-Vietnamese Echinacanthus species. These genes will lead to understanding the adaptation of Echinacanthus species to karst environment. The study will help to resolve the phylogenetic relationship and understand the adaptive evolution of Echinacanthus. It will also provide genomic resources and potential markers suitable for future species identification and speciation studies of the genus.

Dataset Information

Complete chloroplast genome of Oncidium Gower Ramsey and evaluation of molecular markers for identification and breeding in Oncidiinae.

Publications

Complete chloroplast genome of Oncidium Gower Ramsey and evaluation of molecular markers for identification and breeding in Oncidiinae.

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