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Full-length identification and application of wingless gene of Periplaneta americana

A Periplaneta americana, full-length technology, applied in the fields of application, genetic engineering, plant genetic improvement, etc., can solve the problem of little known function

Active Publication Date: 2017-06-16
JIANGSU OPEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, little is known about the role and function of wingless genes during embryonic and post-embryonic development in Periplaneta americana

Method used

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  • Full-length identification and application of wingless gene of Periplaneta americana
  • Full-length identification and application of wingless gene of Periplaneta americana
  • Full-length identification and application of wingless gene of Periplaneta americana

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0073] In Example 1, the full length of the wingless gene of Periplaneta americana was cloned. The specific steps are as follows:

[0074] 1.1 Extract total RNA from tissue samples:

[0075] Extract the total RNA of Periplaneta americana ovum pods, early larvae, late larvae and male adult chest tissue samples, and the extraction of its total RNA is operated on a biological safety cabinet according to the RNeasy Mini Kit (Qiagen company) instructions; then, the total RNA of extraction is dissolved In RNase-free water, the integrity was detected by agarose gel electrophoresis and the purity and concentration were determined by a nucleic acid and protein quantitative detector, and then stored in a -70°C refrigerator for later use.

[0076] The results of electrophoresis were as follows figure 1 Shown: It can be seen from the electrophoresis picture that two obvious bands are 28s and 18s respectively, indicating that the extracted RNA is complete and not degraded, and can be use...

Embodiment 2

[0112] In Example 2, sequence analysis was performed on the full-length cDNA sequence of the Periplaneta americana wingless (PaWg) gene obtained in Example 1.

[0113] The full-length cDNA sequence of PaWg gene was verified by the BLAST program of NCBI. Use ORFFinder (www.ncbi.nlm.nih.gov / gorf / ) to search the open reading frame (open reading frame, ORF) of PaWg cDNA and translate the corresponding amino acid sequence.

[0114] The total length of the gene sequence is 1,456bp, and the predicted open reading frame of the sequence contains 1,194 bases, encoding 397 amino acids, the length of 5'UTR is 157bp, and the length of 3'UTR is 105bp. The start codon of translation is ATG, the stop codon is TGA (positioned at 1349-1351), followed by the tailing signal AATAAA ( Figure 5 ). After the deduced amino acid sequence was analyzed by Blastx homology comparison, it was found that the PaWg gene obtained in this experiment shared 68-76% homology with wingless genes of other species ...

Embodiment 3

[0120] In this embodiment 3, the basic properties of the PaWg protein were analyzed:

[0121] 3.1 Analysis of the physicochemical properties of PaWg protein:

[0122] Using ProtParam tool (http: / / us.expasy.org / tools / protparam.html) online tool to analyze the physicochemical properties of PaWg protein (Gasteiger, E., Hoogland, C., Gattiker, A., Wilkins, M.R., Appel , R.D. and Bairoch, A. 2005. Protein identification and analysis toolson the ExPASy server. Methods Mol Biol 112:531-552), using the Compute pI / Mw online tool (http: / / www.expasy.org / tools / pi_tool. html) to predict the molecular weight and isoelectric point, and sequentially give the number of amino acids (Number of amino acids), molecular weight (Molecular weight), theoretical isoelectric point (Theoretical pI), amino acid composition (Amino acid composition) of PaWg protein , Total number of positively / negatively charged residues, Total number of atoms, Atomic composition, Formula, Instability index, Aliphatic inde...

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Abstract

The invention provides a full-length cDNA of the wingless gene of Periplaneta americana and an encoded protein and cloning method thereof, and application thereof. The full-length cDNA of the wingless gene of Periplaneta americana is named as PaWg, with an accession number of KJ680328 in the GeneBank. The full-length sequence of the cDNA has a length of 1, 456 bp; the open reading frame (ORF) of the cDNA contains 1, 194 bases, coding 397 amino acids; and the nucleotide sequence of the cDNA is as shown in SEQ ID No. 1 in a sequence table. The amino acid sequence of the protein coded by the gene sequence is as shown in SEQ ID No. 2 in the sequence table. The protein has a theoretical isoelectric point PI of 9.46 and a molecular weight of 4.4 kD and is a stable hydrophilic protein. The segment, composed of amino acids from site 1 to site 21, of the amino acid sequence of the protein is a segment of signal peptide; an enzyme site is located between site 21 and site 22; and the protein may contain two N-glycosylation sites, respectively located on asparagine at site 107 and site 343. The invention provides application of the gene, the protein and the cloning method to preparation of cockroach pesticides and biological probes and lays a foundation for research and application related to the wingless gene of Periplaneta americana.

Description

technical field [0001] The invention relates to the cloning and functional identification of a gene, in particular to the full-length cloning, functional identification and application of the Periplaneta americana wingless gene, and belongs to the field of biotechnology. Background technique [0002] Periplaneta americana is the most common cockroach in southern China. It can enter indoors and cause harm. It is one of the dominant hygienic pests. They can be found anywhere indoors as evidenced by dirty mattresses, sheets, upholstered furniture, carpets and they are active in the kitchen [1] . At present, chemical control is still an important means of cockroach control. With the extensive use of chemical insecticides, cockroaches quickly developed resistance and became more and more serious [2] . At the same time, due to the use of chemical pesticides, it will have a serious impact and damage to the ecological environment. Therefore, the use of molecular biological info...

Claims

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Application Information

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IPC IPC(8): C12N15/12C07K14/435C12N15/10C12Q1/68C12N15/11A01N37/46A01P7/04
CPCA01N37/46C07K14/43563C12N15/1096C12Q1/6876C12Q2600/158C12Q2531/113C12Q2549/119
Inventor 陈婉胡超超
Owner JIANGSU OPEN UNIV
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