KR-20260067416-A - Novel Smilax sieboldii chlorosis virus 1 gene from Smilax sieboldii and primer set composition specific to thereof for detecting Smilax sieboldii chlorosis virus 1

Abstract

The present invention relates to a novel Smilax sieboldii chlorosis virus 1 gene derived from Smilax sieboldii and a primer set composition for detecting Smilax sieboldii chlorosis virus 1 specific thereto. Since the primer set of the present invention can specifically detect the novel Smilax sieboldii chlorosis virus, it can contribute to the supply of disease-free plants and quality improvement by diagnosing the pattern of chlorosis virus disease occurrence and identifying abnormal symptoms in Smilax genus plants, including Smilax sieboldii.

Inventors

• 이다현
• 김준혁
• 이재현
• 김현민
• 나채선
• 제상훈

Assignees

• 한국수목원정원관리원

Dates

Publication Date

20260513

Application Date

20241104

Claims (6)

1. Smilax sieboldii chlorosis virus 1 gene consisting of the nucleotide sequence of SEQ No. 1 derived from Smilax sieboldii.
2. A primer set composition for detecting *Clematis chinensis* chlorosis virus 1, comprising one or more oligonucleotide primer sets selected from the group consisting of the oligonucleotide primer sets of SEQ ID NOs. 2 and 3; and the oligonucleotide primer sets of SEQ ID NOs. 4 and 5.
3. A kit for detecting *Clematis chinensis* chlorosis virus 1, comprising an oligonucleotide primer set composition according to claim 2; and a reagent for performing an amplification reaction.
4. In paragraph 3, the kit comprises a reagent for performing the amplification reaction, the reagent comprising DNA polymerase, dNTPs, and a buffer.
5. Step of isolating total RNA from samples suspected of being infected with the blue thorn chlorosis virus; A step of amplifying a target sequence by performing RT-PCR (Reverse Transcription Polymerase Chain Reaction) using the total RNA isolated above as a template and the primer set composition according to claim 2; and A method for detecting the blue thorn chlorosis virus 1, comprising the step of detecting the product of the amplification step.
6. A method according to claim 5, wherein the detection of the amplified product is performed via gel electrophoresis, radiometric measurement, fluorescence measurement, phosphorescence measurement, or DNA chip.

Description

Novel Smilax sieboldii chlorosis virus 1 gene from Smilax sieboldii and primer set composition specific to thereof for detecting Smilax sieboldii chlorosis virus 1 The present invention relates to a novel blue-stemmed rhizome chlorosis virus 1 gene derived from blue-stemmed rhizome and a primer set composition for detecting blue-stemmed rhizome chlorosis virus 1 specific thereto. The emergence and risk of novel and sudden viral diseases are gradually increasing due to ecosystem changes caused by global warming, the evolution and mutation of plants and viruses, and the expansion of agricultural trade. Approximately 1,000 species of plant viruses have been reported worldwide to date, and new viruses continue to be reported in various plants. Currently, about 100 species of viruses have been reported in various plants in Korea. However, detailed investigations into the viruses occurring have been conducted only on a limited number of crops, while only fragmentary investigations have been carried out on most crops. Consequently, the specific details regarding which viral diseases are occurring in many major crops have not yet been clearly identified. Plant disease refers to a condition in which abnormal changes occur in the morphology, physiology, etc., resulting from dysfunction of host cells and tissues caused by the continuous influence of pathogens or environmental factors on a plant host. Causes of such plant diseases include fungi, bacteria, nematodes, mycoplasmas, protozoa, and viruses. However, since viruses are pathogens smaller than 1 µm in size, their physical form cannot be observed even with an optical microscope and requires an electron microscope; therefore, it is difficult to diagnose viruses occurring in plants with the naked eye. Accordingly, various methods for diagnosing viruses have been developed and are being used. Accordingly, in the present invention, a novel viral gene that causes chlorosis in Smilax sieboldii Miq was isolated and identified, and a primer set capable of specifically detecting the novel virus was developed. The aforementioned Smilax china is a climbing shrub belonging to the genus Smilax of the family Smilacaceae. Its stems are green and feature ridges and straight branches. The leaves are ovate-elliptical, hairless, with wavy margins and pointed tips; a pair of tendrils, modified stipules, are located in the center of the petiole. The flowers are yellowish-green and bloom in umbels from May to June, while the fruit is a round berry that matures to black from September to October. In traditional Korean medicine, it is called Jeomeosu (sticky fish whiskers). Its roots and rhizomes are used as medicine and are known to be effective for pain, arthritis, liver cirrhosis, fatty liver, and skin diseases. The young leaves are edible, and the plant grows in forests, distributed in Korea, Japan, China, and other regions. Furthermore, chlorosis is a phenomenon in which green leaves turn yellow due to a lack of chlorophyll, and it occurs as a result of chlorophyll deficiency. Since this chlorophyll deficiency generally occurs when there are not enough nutrients to synthesize all the chlorophyll required by the leaves, if plants affected by chlorosis are not treated, they may eventually die. To date, no viral infections causing chlorosis in the leaves of the Chinese quince have been identified. Therefore, research is needed on viruses that induce chlorosis in Chinese quince plants and methods to detect them. Meanwhile, Korean registered patent No. 2096635 discloses 'a primer set for specifically detecting blackberry chlorosis circular spot virus, a quarantine plant virus, and the use thereof,' and Korean registered patent No. 1024113 discloses 'a PCR primer pair for detecting blackberry chlorosis spot virus and a method for detecting blackberry chlorosis spot virus using the same,' but there is no description of the novel blackberry chlorosis virus 1 gene derived from blackberry and the primer set composition for detecting blackberry chlorosis virus 1 specific thereto according to the present invention. Figure 1 is a photograph showing the symptoms of chlorosis virus in a Smilax sieboldii plant. Figure 2 is a schematic diagram of the whole genome of the novel Smilax sieboldii chlorosis associated virus 1 (SsCAV1) isolated and identified in the present invention. Figure 3 shows the results of an phylogenetic relationship analysis between the novel SsCAV1 of the present invention and 33 types of Carlavirus viruses (Table 3). A is the result of an phylogenetic relationship analysis through homology analysis of the replicase amino acid (aa) sequence, and B is the result of an phylogenetic relationship analysis through homology analysis of the coat protein coding gene (nucleotide, nt) sequence. Figure 4 shows the results of a specificity analysis performed on 12 types of viruses of the genus *Calavirus* using a primer set (Table 4) specific to the novel *Calavirus chlorosis virus* 1 of