Cynoglossus semilaevis sex chromosome-linked DNA segments and application thereof
A semi-smooth tongue sole, chromosome technology, applied in DNA/RNA fragments, recombinant DNA technology, microbial determination/inspection, etc., can solve the problems of high requirements for experimental equipment and reagents, increased costs, long operation time, etc., to save detection time and Cost, increased female ratio, effect of shortened time
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Embodiment 1
[0020] Example 1 Screening of Sex Chromosome-linked DNA Fragments of Semi-smooth Tongue Sole
[0021] The sex chromosome-linked DNA fragment sequence used in the present invention is derived from the genome sequencing results of the half-smooth tongue sole completed by the Aquatic Genome and Cell Engineering Laboratory of the Yellow Sea Fisheries Research Institute of the Chinese Academy of Fishery Sciences. Bioinformatics analysis of the half-smooth tongue sole genome sequence screened out two Z, W chromosome homologous differential DNA fragments. Multiple pairs of primers were designed, and a pair of primers (SEQZW:1, SEQZW:2) with stable results were screened out.
[0022] Select the DNA of female and male half smooth tongue sole with known genetic sex, and use primers SEQZW: 1 and SEQZW: 2 for PCR amplification. The reaction conditions and procedures are as follows: PCR reaction system includes 35.5 μL ddH2O; 5.0 μL 10× Buffer; 4.0 μL dNTP (2.5 mmol / L); 1.0 μL SEQZW:1 (1...
Embodiment 2
[0023] Example 2: Application of Sex Chromosomal Linked DNA Fragment of Semi-smooth Tongue Sole
[0024] 1. Genetic sex identification of tongue sole under laboratory conditions
[0025] The sex chromosome-linked DNA fragment primers SEQZW:1 and SEQZW:2 of tongue sole were used to detect the genetic sex of tongue sole by PCR method. The PCR reaction system is 15 μL, including 1.5 μL of 10×buffer, 0.8 μL of dNTP, 0.3 μL of upstream and downstream primers, 1 μL of template DNA (50 ng / μL), 11.0 μL of ddH2O, and 0.1 μL of rTaq enzyme. PCR amplification program: 10min at 95°C; 35 cycles of 95°C for 30s, 60°C for 30s, and 72°C for 30s; 10min at 72°C, and store at 4°C. Add 10×LoadingBuffer2.0μL, 1.5% agarose gel electrophoresis, 150V, 20 minutes, gel imaging can distinguish genetic gender. ZW females contain two DNA bands of 366bp and 253bp, and ZZ males only have one DNA band of 366bp, such as figure 2 shown.
[0026] 2. Genetic sex identification and pseudo-male screening of h...
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