Watermelon sticky seed gene SNP molecular marker and its screening method and application
A molecular marker, watermelon technology, applied in biochemical equipment and methods, recombinant DNA technology, DNA/RNA fragments, etc., can solve the problem of not obtaining molecular markers, improve the accuracy and selection efficiency, accelerate the improvement process, scientific The effect of practical detection and identification technology
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Embodiment 1
[0034] Example 1: Determination of the sticky seed gene of watermelon seeds and the development of molecular markers
[0035] Simplified genome sequencing of different types of watermelon materials, combined with differences in population polymorphisms to determine the selected regions between populations, and obtaining chromosomal intervals related to sticky-seeded watermelon types, using deep resequencing of sticky-seeded watermelon materials and common watermelon materials The SNP sites in the target chromosome interval were screened, and the CAPS / dCAPS molecular marker primers were designed, and then the CAPS / dCAPS molecular marker primers and watermelon genomic DNA were used for PCR amplification, and finally the amplified products were digested and verified by electrophoresis.
[0036] Specific steps are as follows:
[0037] (1) Choose different types of watermelon materials, including sticky watermelon, common watermelon and wild watermelon;
[0038] (2) Using the meth...
Embodiment 2
[0044] Example 2: Watermelon DNA Molecular Marker Analysis
[0045] 1. Select 5 parts of sticky watermelon, 5 parts of ordinary watermelon and 5 parts of wild watermelon, and use the CTAB method to extract the total DNA of watermelon leaves. Three parallel experiments are set up for each sample. The specific steps are as follows:
[0046] (1) Put 1 g of fresh leaves into a mortar, add liquid nitrogen to grind into powder, then transfer to a centrifuge tube with 1 ml of CTAB extraction solution, mix the two thoroughly, and then place in a constant temperature water bath at 65°C 60 min, during which the mixture was inverted 2 to 3 times;
[0047] (2) After taking it out from the water bath, centrifuge at 8000 rpm for 1 min;
[0048] (3) Take the supernatant and put it in another centrifuge tube, add an equal volume of chloroform: isoamyl alcohol (24:1, V / V), invert gently to mix well;
[0049] (4) Centrifuge at 10,000 rpm for 5 min, and take the supernatant;
[0050](5) Add 0...
Embodiment 3
[0071] Embodiment three: identify and assist the germplasm type of screening watermelon
[0072] Carry out Npr372 molecular marker in 5 parts of sticky watermelon, 5 parts of wild watermelon and 5 parts of common watermelon genotype identification after carrying out embodiment 2, check the distribution situation of two genotypes of Npr372 molecular marker in 93 different types of watermelon materials .
[0073] Table 1 Identification and verification of NPr372 in 93 different types of watermelon materials
[0074] material code breed name Germplasm type NPr372 S001 PI 248178 Sticky Seed Watermelon A S002 PI 494527 Sticky Seed Watermelon A S003 PI 559992 Sticky Seed Watermelon A S004 PI 559993 Sticky Seed Watermelon A S005 PI 559994 Sticky Seed Watermelon A S006 PI 559996 Sticky Seed Watermelon A S007 PI 559999 Sticky Seed Watermelon A S008 PI 560000 Sticky Seed Watermelon A S009 PI 56...
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