A SNP molecular marker affecting the number of thoracic vertebrae in sheep and its application
A molecular marker and molecular marker-assisted technology, applied in the field of molecular biotechnology and molecular markers, to achieve the effects of improving reproductive performance, rapid and accurate selection and improvement, and increasing core competitiveness
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Embodiment 1
[0036] 1. Experimental animals
[0037] The experimental sheep group used in the present invention is 496 Ujimqin sheep selected from the grassland area of East Ujimqin Banner in Inner Mongolia Autonomous Region.
[0038] 2. Sample collection
[0039] The muscle tissue of the slaughtered sheep was collected and soaked in ethanol with a volume fraction of 75%, and stored in a -20°C refrigerator for later use.
[0040] 3. Whole sheep genome 10× resequencing
[0041] Using the muscle tissue collected from each of the 496 sheep in step 1, the whole genome DNA was extracted by the standard phenol-chloroform method, and the DNA concentration and OD ratio of each sample were accurately determined by the Nanodrop2000 / 2000C nucleic acid and protein detector (OD260 / 280, OD260 / 230). Qualified DNA samples detected by the NanoDrop2000 / 2000C Nucleic Acid Protein Detector. Pipette 3 μL of the extracted DNA sample to be tested and mix it with 1.5 μL Loading Buffer, load the sample onto ...
Embodiment 2
[0051] Example 2 Target DNA sequence amplification and sequencing
[0052] (1) Primer design
[0053] The DNA sequence of SEQ ID NO: 1 on chromosome 7 of sheep was downloaded from the UCSC website (http: / / genome.ucsc.edu / ). And use primer design software primer premier 3.0 to design primers.
[0054] The DNA sequences of the designed primers are as follows:
[0055] P001-F: 5'-TGCAATTGGCTGTTTGAGTC-3';
[0056] P002-R: 5'-ACTTGCATCCCTTCAGCAGT-3';
[0057] (2) PCR amplification
[0058] Add 1 μL of DNA template, 10.5 μL of double distilled water, 12.5 μL of 2×Dream Taq Green PCRMaster mix (Thermofisher), and 0.5 μL of primers P001-F and P002-R into the 25 μL reaction system.
[0059] The PCR reaction conditions were: pre-denaturation at 95°C for 5 min; 30 cycles of denaturation at 95°C for 30 s, annealing at 60°C for 30 s, and extension at 72°C for 30 s; and extension at 72°C for 10 min.
[0060] (3) DNA sequence determination
[0061] DNA sequence sequencing identificati...
Embodiment 3
[0064] Example 3 SNP site 82509619G>A effect analysis of molecular markers
[0065] Through molecular marker-assisted selection and elimination of sheep with genotypes of GA and GG in the group, we have significantly increased the total number of thoracic vertebrae in the group, promoted the breeding process of sheep, increased the meat production performance of the sheep group, and driven the economy of the mutton sheep industry Benefits are maximized.
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