Watermelon reticulate pattern peel overlay characteristic SNP site, CAPS molecular marker, primer and application thereof
A molecular marker, watermelon technology, applied in its CAPS molecular marker, watermelon reticulated peel characteristic SNP loci, can solve the lack of such molecular markers and other problems, to achieve high specificity effect
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Embodiment 1
[0023] Example 1 Acquisition of characteristic SNP loci of watermelon reticulated peel
[0024] The present invention takes 1022 watermelon germplasm resources selected by the "National Watermelon and Melon Mid-term Library" of the Zhengzhou Fruit Tree Research Institute of the Chinese Academy of Agricultural Sciences as samples, and uses the Illumina HiSeq4000 sequencer to perform simplified genome sequencing to obtain 142.19 Gb of data, which covers the average coverage of the reference genome. The accuracy was 3.54%, and the average sequencing depth of the covered region was 6.3X. Finally, 121,405 SNP marker sites were obtained through identification screening. The 1022 watermelon resources were divided into two categories according to the characteristics of pericarp covering: whether the reticulation was divided into two categories as phenotypes for GWAS (genome-wide association analysis), and a candidate region related to the characteristics of watermelon pericarp was obt...
Embodiment 2
[0025] Example 2 Design and identification of CAPS marker primers for reticulate peel features
[0026] Combined with the deep resequencing results of some resources, the SNP and InDel sites near the peak in the candidate region were mined, and co-dominant molecular markers were developed to verify in the resources, and finally the method for detecting the characteristics of the watermelon reticulate skin covered by the present invention was obtained. CAPS marker, and its primer sequence is as follows (see SEQ ID NO.1, SEQ ID NO.2 in Sequence Listing):
[0027] Upstream primer: 5'-TGGACTTCTAAATCGGATTCTC-3'
[0028] Downstream primer: 5'-TTCTTAACGGTTTCTTCTTGGT-3'.
[0029] Utilize the above-mentioned molecular markers to identify the method of watermelon reticulate peel feature, which mainly includes the following steps:
[0030] (1) The total DNA of leaves was extracted by CTAB method. The specific steps are as follows:
[0031] Take 1 g of fresh leaves into a mortar, ad...
Embodiment 3
[0074] Example 3 Identification and verification of CAPS markers in watermelon germplasm
[0075] The application of CAPS markers in the identification of watermelon peel features, the main steps are as follows:
[0076] (1) Using the CAPS molecular markers and method steps described in Example 2, 62 watermelon germplasms with known pericarp striae characteristics were genotyped.
[0077] (2) According to the electrophoresis results of the enzyme digestion products, if only a band of 177bp is obtained, the material to be tested is characterized by reticulate pericarp; if only two bands of 82bp and 95bp are obtained, the material to be tested is non-reticulated. The characteristics of the pericarp covering pattern; if three bands of 82bp, 95p and 177bp are obtained, the material to be tested exhibits the characteristics of non-reticulate pericarp covering; see image 3 .
[0078] (3) Identification results
[0079] The detection results of 62 watermelon germplasms showed tha...
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