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Molecular marker primer and method used for identification of prunus pedunculata, prunus mongolica, and prunus triloba

A long-stalked almond, molecular marker technology, applied in biochemical equipment and methods, microbiological determination/inspection, DNA/RNA fragments, etc., can solve problems such as complex structure and composition, gene rearrangement, repeat loss, etc., to improve Detecting the Effects of Speed ​​and Efficiency

Active Publication Date: 2018-12-14
INNER MONGOLIA AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Among the plant DNA barcodes, available genomic DNAs include nuclear genomic DNA (nrDNA), chloroplast genomic DNA (cpDNA) and mitochondrial genomic DNA (mtDNA). Among them, nrDNA has the fastest evolution rate and is a fully autonomous genetic system in the nucleus. The structure and composition are extremely complex, each gene family forms a functional regulatory network, and many genes have multiple copies, and there are problems such as orthologs and paralogs
However, mtDNA is prone to gene rearrangement, duplication and loss, and the copy number of plant mtDNA is low, so it is difficult to extract and purify. Therefore, the application range of nrDNA and mtDNA in plant identification is limited.

Method used

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  • Molecular marker primer and method used for identification of prunus pedunculata, prunus mongolica, and prunus triloba
  • Molecular marker primer and method used for identification of prunus pedunculata, prunus mongolica, and prunus triloba
  • Molecular marker primer and method used for identification of prunus pedunculata, prunus mongolica, and prunus triloba

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] 1. Collection of Leaf Samples

[0053] The samples were identified and collected by the Inner Mongolia Forest Tree Breeding Center, and 5 fresh, healthy, and pest-free leaf samples were collected from different origins of almonds, Mongolian almonds, and eucalyptus. As described in Table 1.

[0054] Table 1 Leaf samples and sources of almonds, Mongolian almonds and Yuyemei

[0055]

[0056] 2. Extraction of total genomic DNA from leaf samples

[0057] The total genomic DNA of the leaf sample was extracted by the plant DNA rapid extraction kit, and its quality was detected based on 1.2% agarose gel electrophoresis, and the results showed as follows: figure 1 As mentioned above, the brightness and uniformity of the electrophoresis strips and the light spots of the sample port are all good, indicating that the samples are of high purity and no pollution, and can be directly used in subsequent molecular biology experiments.

[0058] 3. PCR amplification for the target ...

Embodiment 2

[0074] Embodiment 2 result analysis

[0075] After the PCR product was purified, two-way sequencing was performed, and the sequences of the leaf sample group of Long handle almond, Mongolian almond leaf sample group and Yuyemei leaf sample group were compared. The result of the alignment of the trnK-UUU fragment sequence of the chloroplast gene with the sample group of Yuyemei leaves is as follows:

[0076] Almonds with long handles:

[0077] ACTCGAACCCGGAACTAGTCGGATGGAGTAGATAATTTCCTTGATTAGAAAATTTTAAATAAAATAGGGAAAAA(A) + CCCCTCCCCAAACCGTGCTTGCATTTTTCATTGCACACGGCTTTCCCTATGTATACATCTAAAATTCAGTCCCTTCCCTATACACGACTCTAAGAAAGTTGAATACTCAGTTGATCGACCCTCAATCTTACTGTATGAACATTTCATAATAGAAATAAATGCAAATTTTTT(ATAGAAATAAATGCAAATTTTTT) + GTTATCTCTTTCATTATTTAAAGA G AATTCCATTTCTACGATCGCATAACCAATTATTCATAATTGATTAGATCATTGATGCAAAAAATA T CCAAATACCAAATCCGACCTCTATAAAATTTCTTAAAAGTAAAAGTATAAGAAGCTCTTGGGAAGACCAA A GAAAGAATCTGTTCTTCTTCCGTAAAGAATTCTTCCAATAATTTAGAACCTAATCTTTTCAAAAAAGTTCGTACAGTACTTTTTGTGTTTAC...

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Abstract

The invention discloses a molecular marker primer and a method which are used for identification of prunus pedunculata, prunus mongolica, and prunus triloba. The molecular marker primer comprises a forward primer and a reverse primer; the sequence of the forward primer tKF is ACTCGAACCCGGAACTAG, and the sequence of the reverse primer tKR is CGACCGATTTCTGCGTAT. The identification method comprises following steps: 1, leaf sample collecting; 2, leaf sample total genome DNA extraction; 3, amplification of target genes in leaf sample total genome DNA obtained in step 2 using the molecular marker primer used for identification of prunus pedunculata, prunus mongolica, and prunus triloba, and obtaining of amplification products; 4, purifying and sequencing of the amplification products obtained instep 3; and 5, comparison analysis on sequencing results obtained in step 4. According to the method, trnK-UUU intergenic region based base difference can be taken as the molecular marker primer usedfor identification of prunus pedunculata, prunus mongolica, and prunus triloba germplasm distinguishing, the identification method is simple, and operation is convenient.

Description

technical field [0001] The present invention relates to the field of biotechnology. Specifically, it relates to a molecular marker primer and a method for identifying long-handled almonds, Mongolian almonds and elm leaf plums. Background technique [0002] Long handle almond (Prunus pedunculata Pall), Mongolian almond (Prunus mongolica Maxim) and elm leaf plum (Prunus triloba Lindl) are unique economic forest tree species in my country. Extremely cold and drought resistant, it is the preferred tree species for ecological construction in semi-arid desertification areas in my country. After research, it was found that the oil content of long-stalked almond seeds is about 56%, which is higher than that of Mongolian almonds and elm plums. Its oil is rich in unsaturated fatty acids (96%-98%), ranking first among all vegetable oils. More than 70%, it belongs to high oleic vegetable oil, which is equivalent to olive oil known as "the queen of edible vegetable oil", and is a functi...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895C12Q2600/13C12Q2600/156
Inventor 包文泉敖敦
Owner INNER MONGOLIA AGRICULTURAL UNIVERSITY
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