Construction of whole genome full-length cDNA FOX-hunting overexpression library in Brassica napus

doi:10.19802/j.issn.1007-9084.2020044

Abstract

Abstract:

Genome-wide overexpression library is an important basis for systematic characterization of gene function in Brassica napus. Full-length cDNA FOX-hunting overexpression library was constructed by combining FOX-hunting and Gateway recombination techniques with full-length cDNA of mixed samples of seedling roots and leaves treated with 4 hormones and 4 abiotic stresses, and mature seeds and silique pericarps in B. napus. Titer of the primary library was 3.0×106, and total CFU was 1.2×107. The secondary FOX-hunting overexpression library had a titer of 2.6×106 and a total CFU of 1.04×107. PCR identification showed that the positive clone rate was 96%, and the amplificons were all exhibited as single band random distributed within 250-3000 bp, indicating that the primary library had been well recombined into pJVC55 vector. Sequencing results of Agrobacterium-transformed clones showed that 12 of the 140 clones were non-specific. GO enrichment analysis found that, except for 21 genes
enriched in photosynthesis, the remaining genes randomly distributed in transcription factors and kinases, and their functions were involved in multiple aspects involved organ development, metabolism and stress response. The results showed that our FOX-hunting overexpression library of B. napus had rich polymorphism and high integrity,but no functional preference, which laid an important foundation for the functional genome study in B. napus.

Key words: , Brassica napus；full-length cDNA；FOX-hunting；overexpression；library

CLC Number:

中图分类号：S565.4

ZHANG Kai, WEI Si-yu§, CHANG Wei, FAN Yong-hai, LU Kun. Construction of whole genome full-length cDNA FOX-hunting overexpression library in Brassica napus[J]. CHINESE JOURNAL OF OIL CROP SCIENCES, 2021, 43(3): 435-.

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