Cloning and VIGS Functional Confirming of NBS Type Diseaseresistance Gene from Cucurbita ficifolia
CSTR:
Author:
Affiliation:

Clc Number:

Fund Project:

  • Article
  • |
  • Figures
  • |
  • Metrics
  • |
  • Reference
  • |
  • Related
  • |
  • Cited by
  • |
  • Materials
  • |
  • Comments
    Abstract:

    Cucurbita ficifolia Bouche. is a unique cucurbits germplasm resource in Yunnan Province with highly Fusarium wiltresistance genetic trait. To explore the function of NBSLRR type genes from C. ficifolia, in this study, we isolated the NBS type gene of CfRFN2 (GenBank ID:MK618462). We obtained a full length 4 303 bp by divided three fragment cloning using RTPCR method from leaves of C. ficifolia. The CfRFN2 gene was annotated as a homologous gene of transcriptionoid X1 of Arabidopsis diseaseresistant protein At4g27190, with complete coding frame length of 4 092 bp and encoding 1 363 amino acids, and its deduced protein contained one NBARC conserved domain and two LRR domains. The CfRFN2 gene belongs to a hydrophilic protein containing the signal peptide. The results of nucleotide similarity analysis indicated that the identity of CfRFN2 gene varied from 87% to 98% when compared with the other disease resistance genes in Cucurbitaceae crops. Phylogenetic analysis results showed that CfRFN2 protein and other NBS type protein from cucurbits crops were clusterd into one branch, indicating it had closer genetic relation among cucurbits crops than with other species NBS protein. Moreover, CfRFN2 had the closest genetic relation with RPS2 of Cucurbita pepo and Cucurbita moschata, Cucurbita maxima RPS2like protein, followed by Cucurbita sativus At4g27190 and Momordica charartia At4g27220, while relatively quite far from Cucurbita melo At4g27190. The tissue specific expression results indicated that the CfRFN2 gene was highly expressed in leaves than that in stems, and had low expression level in fruits and roots. Furthermore, based on the tobacco rattle virus vector system, we also constructed VIGS silencing vector pTRV2CfRFN2, constantly the seedlings of C. ficifolia were infected by Agrobacterium tumefaciens carrying silencing vector and inoculated with Fusarium oxysporum after 4 weeks infection (wild type plants inoculation pathogeny simultaneously). The qRTPCR analysis showed that, compared to the wild type plant, the expression of CfRFN2 gene in silencing plants decreased by 34.75% and 98.27%, respectively after 2 and 4 days inoculation. Moreover, the average disease index of silencing plants was 1.32 times that of wild type plants. This preliminarily results suggested that CfRFN2 gene in C. ficifolia had the function of resistance to Fusarium wilt. The cloning and VIGS confirmation of NBS type gene CfRFN2 would be the basis for digging more disease resistant genes in C. ficifolia, and thus provide a new clue for carrying out molecular breeding and exploring the disease resistant mechanism in cucurbits crops.

    Reference
    Related
    Cited by
Get Citation

DING Yumei, HOU Siming, YAO Chunxin, YANG Jiazhen, XIE Junjun, ZENG Yawen, YANG Zhengan. Cloning and VIGS Functional Confirming of NBS Type Diseaseresistance Gene from Cucurbita ficifolia[J]. Acta Botanica Boreali-Occidentalia Sinica,2021,41(5):727-737

Copy
Share
Article Metrics
  • Abstract:
  • PDF:
  • HTML:
  • Cited by:
History
  • Received:
  • Revised:
  • Adopted:
  • Online: June 22,2021
  • Published:
Article QR Code