Encoding gene from anopheles sinensis aquaporin 2 and application of anopheles sinensis aquaporin 2
An aquaporin and encoding gene technology, applied to the encoding gene and application of Anopheles sinensis aquaporin 2 (AsAQP2), aquaporin and its encoding gene and application field, can solve research problems and achieve efficient water transport Function, the effect of good application prospects
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0022] Example 1 Cloning of Anopheles sinensis aquaporin 2 cDNA sequence
[0023] According to the conserved amino acid sequence of aquaporin in Anopheles gambiae, the demerger primers were designed, and the full-length cDNA of this gene was cloned from Anopheles sinensis by RT-PCR combined with RACE technology. After sequence comparison, the gene has a high homology with one of the aquaporins in Anopheles gambiae. The expression abundance of this protein in Anopheles gambiae is high, and it has a very efficient water transport function. Design specific primers based on sequence information:
[0024] 3F: 5'-ATGACTGAAA GCGCAGGAGT G-3',
[0025] 3R: 5'-TTAAAAATCG TATGAATTTG ATTCTTCG-3',
[0026] Using the adult Anopheles sinensis mRNA, the cDNA template was obtained using Invitrogen’s SuperScriptⅢ reverse transcription kit, and the PCR reaction was prepared according to the following reaction system:
[0027] cDNA template 2 μL Upstream primer 3F (20 μM) 0....
Embodiment 2
[0031] Example 2 In vitro function identification of Anopheles sinensis aquaporin 2 (Xenopus oocyte system)
[0032] Primers were designed according to the coding region of the gene sequence, and BglⅡ restriction sites were added at both ends. The specific primer sequences are as follows:
[0033] AQP2BglⅡF0': 5'-AGA agatct ATGACTGAAA GCGCAGGAGT G-3',
[0034] AQP2BglⅡR0': 5'-AGA agatctTTAAAATCGT ATGAATTTGA TTC-3',
[0035] The endonuclease BglⅡ digested and recovered the PCR product and the oocyte expression vector pXβG-myc respectively, and ligated them overnight at 16°C with T4 ligase. The constructed plasmid was identified by PCR, enzyme digestion and sequencing respectively, and after confirming that the direction and sequence were correct, the extracted plasmid was linearized. The mMESSEGemMACHINE T3 in vitro transcription kit of Ambion Company was used for in vitro transcription. For details, please refer to the company's instruction manual. The resulting product wa...
Embodiment 3
[0041] Example 3 Expression of Aquaporin 2 in Anopheles sinensis at different developmental stages and in different tissues of adult mosquitoes
[0042] Collect mosquito samples from the same batch of Anopheles sinensis at different developmental stages, namely: mosquito eggs, 4th instar larvae, pupae, adults (male mosquitoes, female mosquitoes); and the collection of different tissues of Anopheles sinensis, from the same batch 50-60 female mosquitoes were taken, and the head, midgut, ovary, and fat body were separated under a dissecting microscope to extract total RNA for fluorescent quantitative PCR detection.
[0043] The collected frozen specimens were ground and homogenized on ice. Total RNA was extracted by Qiagen RNeasy kit and treated with RNase-free DNAse to remove contamination from genomic DNA. Take 2 μg of total RNA and synthesize cDNA by reverse transcription with random primers according to the instructions of the Toyobo reverse transcription kit. cDNA samples ...
PUM
Abstract
Description
Claims
Application Information
- R&D Engineer
- R&D Manager
- IP Professional
- Industry Leading Data Capabilities
- Powerful AI technology
- Patent DNA Extraction
Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.
© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com