Litopenaeus vannamei heat shock protein 40 gene lvHSP40 and application thereof in inhibition of ovary development of litopenaeus vannamei
A heat shock protein and ovarian development technology, applied in the field of aquatic organisms, can solve problems such as difficulty in synchronous development of gonads, lack of ovarian development, influence on mating and reproductive efficiency, etc., and achieve the effect of great economic value and wide application prospects.
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Embodiment 1
[0021] Example 1: Cloning and sequencing of the heat shock protein 40 gene lvHSP40 of Litopenaeus vannamei
[0022] Using the hepatopancreas tissue of Litopenaeus vannamei as material, the total RNA was extracted using the Trizol (purchased from Invitrogen Company) method, the remaining genomic DNA was digested using DNase I (purchased from Invitrogen Company), and PrimeScript was used to TM II kit (purchased from Takara Company) was reverse transcribed into the first strand of cDNA. Based on a 1075-base assembly fragment annotated as HSP40 obtained from the second-generation sequencing of the Litopenaeus vannamei hepatopancreas transcriptome (provided by Huada Genomics) on the Illumina Hiseq platform we constructed, the design gene-specific (provided by Shanghai Shenggong The company synthesized) primers were amplified to obtain the cDNA middle fragment of the heat shock protein 40 gene lvHSP40 of Litopenaeus vannamei. On the basis of obtaining the intermediate fragment, th...
Embodiment 2
[0036] Example 2: Injection of siRNA HSP40-474 inhibits gonad development of female Litopenaeus vannamei
[0037] The siRNA sequences HSP40-42, HSP40-474, and HSP40-825 were designed according to the cDNA sequence of the heat shock protein 40 gene lvHSP40 of Litopenaeus vannamei (as shown in Table 1). The siRNA was synthesized by Shanghai Sangong Company. The mRNA expression of lvHSP40 was knocked down by RNA interference by injection method, and the mRNA expression level of lvHSP40 in the hepatopancreas (Hp) and ovary (Ov) of Litopenaeus vannamei was detected at 12 hours and 24 hours after injection, and it was found that 300ng / g body weight The dose of siRNA HSP40-474 injection had the best knockdown effect on the mRNA expression of Litopenaeus vannamei lvHSP40 at each time point (as shown in Table 1). For siRNA HSP40-474, the nucleotide sequence of its sense strand is shown in SEQ ID NO.3, and the nucleotide sequence of its antisense strand is shown in SEQ ID NO.4.
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