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Fusion gene for three sweet potato viruses and interference carrier thereof

A technology of sweet potato virus and fusion gene, applied in the field of bioengineering

Active Publication Date: 2011-10-05
HENAN ACAD OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] The technical problem to be solved in the present invention is to overcome the shortcomings of the existing prevention and treatment of sweet potato virus diseases, provide a fusion gene for three kinds of sweet potato viruses, and also provide an RNA interference carrier that has obvious interference and inhibition effects on the three viruses

Method used

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  • Fusion gene for three sweet potato viruses and interference carrier thereof
  • Fusion gene for three sweet potato viruses and interference carrier thereof
  • Fusion gene for three sweet potato viruses and interference carrier thereof

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Embodiment 1

[0032] Embodiment 1: Fusion gene and its interference carrier for three kinds of sweet potato viruses

[0033] (1) Materials and methods

[0034] 1. Design and synthesis of fusion genes of SPFMV, SPLV and SPVG

[0035] Analyze the conserved regions of the CP genes of SPFMV, SPLV and SPVG, and select fragments of about 200 bp each, of which the conserved region of the SPFMV CP gene is 198 bp, located at the 748-945 bp of its CP gene; the conserved region of the SPLV CP gene is 189 bp, located at The 691-879bp of its CP gene; the 198bp conserved region of the SPVG CP gene is located at the 868-1065bp of its CP gene. Fusion was carried out according to the sequence of SPFMV-SPLV-SPVG, and the complete target gene was named as SPFLG. In order to facilitate the construction of the vector, the 5' end of the gene was introduced Xba I site, introduced at its 3' end Speech I site, and add three protective bases each to make the enzyme cutting site more effective. The length of t...

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Abstract

The invention relates to a fusion gene for three sweet potato viruses and an interference carrier thereof. The sequence of the fusion gene for three sweet potato viruses is SEQ1 and is named as SPFLG. The construction method of the interference carrier comprises the following steps: designing two pairs of primers, that is, a pair of SPFLG-XbaI and SPFLG-SpeI and a pair of SPFLG-KpnI and SPFLG-ClaI; cloning the SPFLG gene onto a T carrier, carrying out PCR amplification by respectively using the two pairs of primers with the SPFLG-T as a template, respectively carrying out enzyme digestion on the obtained target gene SPFLG with XbaI and SpeI and with KpnI and ClaI, and respectively cloning the digested target gene SPFLGs onto the two sides of an intron of a pBlueNiRIntron carrier in a manner of inverted repeats; carrying out double enzyme digestion with XbaI and KpnI, recovering a segment of the target gene and cloning the segments onto an intermediate carrier pROK219; carrying out enzyme digestion of pROKSPFLG with EcoRI and HindIII, recovering a target segment comprising a 35S promoter, inverted repeats and an NOS terminator, cloning the target segment onto a plant expression vector pBIN19 and naming the vector to be pBINSPFLG. According to results of experiments, RNA interference carrier constructed in the invention has a significant interference effect on the three viruses, namely, SPFMV, SPLV and SPVG, and lays a foundation for cultivation of a transgenic sweet potato with resistance to the three viruses.

Description

[0001] technical field [0002] The invention relates to genes and interference vectors of sweet potato viruses, belonging to the technical field of bioengineering, in particular to fusion genes and interference vectors for three sweet potato viruses. Background technique [0003] Sweet potato is an important grain, feed, industrial raw material and new energy crop. In recent years, the use of sweet potato has attracted more and more attention. Sweet potato virus disease is one of the important limiting factors affecting the production of sweet potato. The infection of virus disease can cause the yield decrease, quality deterioration and species degradation of sweet potato, generally can reduce the yield by 30%-50%. Sweet potato feather mottle virus ( Sweet potato feathery mottle virus , SPFMV), sweet potato latent virus ( Sweet potato latent virus , SPLV) and sweet potato G virus ( Sweet potato virus G , SPVG) are the three main viruses that harm sweet potato, and thes...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/62C12N15/82C12N15/66A01N61/00A01P1/00
Inventor 张振臣秦艳红张德胜张磊乔奇田雨婷郑文明
Owner HENAN ACAD OF AGRI SCI
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