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KR-20260065702-A - SNP MARKER COMPOSITION FOR IDENTIFICATION OF DUCKWEED SPECIES AND METHOD FOR IDENTIFICATION OF DUCKWEED SPECIES USING THE SAME

KR20260065702AKR 20260065702 AKR20260065702 AKR 20260065702AKR-20260065702-A

Abstract

The present invention relates to an SNP marker composition for identifying duckweed species and a method for identifying duckweed species using the same. More specifically, the invention relates to an SNP marker composition capable of identifying duckweed species such as Spirodela polyrhiza, Lemna minor, Wolffia lingulata, Wolffia brasiliensis, Wolffia australiana, Wolffia globosa, Wolffia arrhiza, Lemna turionifera , and Spirodela intermedia , and a method for identifying duckweed species using the same. The present invention identified genetic markers for identifying duckweed species by selecting single nucleotide polymorphisms (SNPs) within the duckweed chloroplast genome using a random forest algorithm.

Inventors

  • 강양제
  • 박하림

Assignees

  • 경상국립대학교산학협력단

Dates

Publication Date
20260511
Application Date
20241101

Claims (7)

  1. (a) a polynucleotide consisting of 5 to 987 consecutive nucleotides including a single nucleotide polymorphism (SNP) marker of the 48th nucleotide of the nucleotide sequence represented by SEQ ID NOs 1 to 9, or a polynucleotide complementary thereof; (b) a polynucleotide consisting of 5 to 987 consecutive nucleotides, including a single nucleotide polymorphism (SNP) marker of the 150th nucleotide of the nucleotide sequences represented by SEQ ID NOs 1 to 9, or a polynucleotide complementary thereof; (c) a polynucleotide consisting of 5 to 987 consecutive nucleotides including a single nucleotide polymorphism (SNP) marker of the 201st nucleotide of the nucleotide sequence represented by SEQ ID NOs 1 to 9, or a polynucleotide complementary thereof; (d) a polynucleotide consisting of 5 to 987 consecutive nucleotides including a single nucleotide polymorphism (SNP) marker of the 204th nucleotide of the nucleotide sequences represented by SEQ ID NOs 1 to 9, or a polynucleotide complementary thereof; (e) a polynucleotide consisting of 5 to 987 consecutive nucleotides including a single nucleotide polymorphism (SNP) marker of the 240th nucleotide of the nucleotide sequences represented by SEQ ID NOs 1 to 9, or a polynucleotide complementary thereof; (f) a polynucleotide consisting of 5 to 987 consecutive nucleotides including a single nucleotide polymorphism (SNP) marker of the 399th nucleotide of the nucleotide sequences represented by SEQ ID NOs 1 to 9, or a polynucleotide complementary thereof; (g) a polynucleotide consisting of 5 to 987 consecutive nucleotides including a single nucleotide polymorphism (SNP) marker of the 438th nucleotide of the nucleotide sequences represented by SEQ ID NOs 1 to 9, or a polynucleotide complementary thereof; (h) a polynucleotide consisting of 5 to 987 consecutive nucleotides, including a single nucleotide polymorphism (SNP) marker of the 744th nucleotide of the nucleotide sequence represented by SEQ ID NOs 1 to 9, or a polynucleotide complementary thereof; (i) a polynucleotide consisting of 5 to 987 consecutive nucleotides including a single nucleotide polymorphism (SNP) marker of the 759th nucleotide of the nucleotide sequences represented by SEQ ID NOs 1 to 9, or a polynucleotide complementary thereof; (j) a polynucleotide consisting of 5 to 987 consecutive nucleotides including a single nucleotide polymorphism (SNP) marker of the 870th nucleotide of the nucleotide sequence represented by SEQ ID NOs 1 to 9, or a polynucleotide complementary thereof; and (k) a polynucleotide consisting of 5 to 987 consecutive nucleotides including a single nucleotide polymorphism (SNP) marker of the 441st nucleotide of the nucleotide sequence represented by SEQ ID NOs 1 to 9, or a polynucleotide complementary thereof; One or more polynucleotides selected from the group consisting of, or comprising a polynucleotide complementary thereof, SNP marker composition for identifying duckweed species.
  2. In paragraph 1, The above duckweed species is An SNP marker composition for identifying duckweed species, characterized by being one or more selected from the group consisting of Spirodela polyrhiza, Lemna minor, Wolffia lingulata, Wolffia brasiliensis, Wolffia australiana, Wolffia globosa, Wolffia arrhiza, Lemna turionifera, and Spirodela intermedia .
  3. A composition for identifying duckweed species, comprising a preparation capable of detecting the SNP marker composition for identifying duckweed species of claim 1.
  4. In paragraph 3, The above preparation is A composition for identifying duckweed species, characterized by having one or more selected from the group consisting of primers and probes.
  5. In paragraph 4, The above primer is A composition for identifying duckweed species, characterized by comprising a forward primer represented by SEQ ID NO. 10 and a reverse primer represented by SEQ ID NO. 11.
  6. A step of amplifying a polynucleotide containing the SNP marker of claim 1 from the DNA of a sample isolated from an individual; and A step of determining the base of the amplified SNP marker site; comprising Method for identifying duckweed species.
  7. In paragraph 6, The above duckweed species is A method for identifying duckweed species, characterized by having one or more selected from the group consisting of Spirodela polyrhiza, Lemna minor, Wolffia lingulata, Wolffia brasiliensis, Wolffia australiana, Wolffia globosa, Wolffia arrhiza, Lemna turionifera, and Spirodela intermedia .

Description

SNP Marker Composition for Identification of Duckweed Species and Method for Identification of Duckweed Species Using the Same The present invention relates to an SNP marker composition for identifying duckweed species and a method for identifying duckweed species using the same. More specifically, the invention relates to an SNP marker composition capable of identifying duckweed species such as Spirodela polyrhiza, Lemna minor, Wolffia lingulata, Wolffia brasiliensis, Wolffia australiana, Wolffia globosa, Wolffia arrhiza, Lemna turionifera , and Spirodela intermedia , and a method for identifying duckweed species using the same. The systematic classification of organisms is the cornerstone of biological research and provides an important framework for identification. Traditionally, species classification has been based on morphological characteristics such as shape, size, and color. However, classifying species based on morphological characteristics suffers from a lack of accuracy due to the classifier's subjective interpretation and changes in the organism's appearance caused by environmental shifts. Furthermore, as research on biodiversity has advanced, it has been revealed that species with similar morphological characteristics may possess different genetic backgrounds, while species with different morphological characteristics may share similar genetic backgrounds. Recently, there has been a growing need for objective and reliable technologies to classify species relationships, and DNA barcoding is a technology developed in response to this situation. DNA barcoding facilitates species identification by utilizing unique variations within the DNA sequences of each biological species. More specifically, PCR products amplified by PCR targeting specific regions of the genome can serve as unique sequence tags for each species. DNA barcodes contain universally used target genes or regions and primers designed for said genes or regions. In animals, the region is COI; in plants, it is matK or rbcL, etc.; and in fungi, it is the ITS region, etc. Many researchers have been able to extract sequence data of only the target gene regions of various species by PCR and sequencing specific parts of the target using designed primers. As this sequence data has accumulated, it has now become possible to classify species by performing PCR with primers. However, even if primers are designed for universality, frequent sequence changes can lead to a failure to react with the target sequence, resulting in a low PCR success rate. Consequently, there is a growing need to develop methods to identify the most suitable DNA barcode for each species and improve the resolution of species classification. The problems with DNA barcoding described above have become even greater issues in taxa such as duckweed. Duckweed is difficult to classify taxonomically, as it includes 38 species across 5 genera. Furthermore, morphological classification is very difficult due to the small leaf size, which is only 1 to 15 mm in diameter or length, and the simple structure. Additionally, whole genome sequences have been found for only 3 species of duckweed, and only 9 species have complete chloroplast genome sequences. Therefore, there is a growing need for a new approach that can more easily identify duckweed species. Figure 1 is a schematic diagram illustrating the process of collecting and preprocessing duckweed chloroplast sequences to select SNPs for duckweed species identification. Figure 2 is a schematic diagram illustrating the process of selecting high-importance SNPs using a random forest algorithm to select SNPs for identifying duckweed species. Figure 3 shows the results of analyzing the characteristic importance values of SNPs using the Random Forest algorithm. Figure 4 shows the results of selecting SNPs in order of containing a large number of SNPs with high characteristic importance values. Figure 5 shows the sequence of the duckweed ndhH gene, SNPs within the sequence, and primers. Figure 6 shows the sequence of the duckweed ndhH gene, SNPs within the sequence, and primers. Figure 7 shows the results of PCR amplification of duckweed genome DNA using primers of SNP marker genes. Figure 8 shows the classification of duckweed species by the final selected SNPs for duckweed species identification. The terms used in this specification have been selected based on currently widely used general terms whenever possible, taking into account their functions in the present invention; however, these terms may vary depending on the intent of those skilled in the art, case law, the emergence of new technologies, etc. Additionally, in specific cases, terms have been arbitrarily selected by the applicant, and in such cases, their meanings will be described in detail in the relevant description of the invention. Therefore, the terms used in this invention should be defined not merely by their names, but based on their meanings and the overall content of