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Journal of Chinese Integrative Medicine ›› 2006, Vol. 4 ›› Issue (6): 639-643.doi: 10.3736/jcim20060620

• Original Experimental Research • Previous Articles     Next Articles

Construction of a plant effective expression vector containing the gene of hepatitis B virus surface antigen

Bing-ying Lin1,2,Zhi-qiang Jin2,Mei Li1   

  1. 1. Xiamen Overseas Chinese Subtropical Plant Introduction Garden, National Plant Introduction Quarantine Base, Xiamen, Fujian Province 361002, China
    2. State Key Laboratory of Tropical Crops Biotechnology, The Chinese Academy of Tropical Agricultural Sciences, Haikou, Hainan Province 571101, China
  • Online:2006-11-30 Published:2006-11-15

Objective

To construct a plant effective expression vector driven by a fruit specific promoter for the expression of hepatitis B virus surface antigen (HBsAg), to further improve the expression of exogenous gene in plant, and to prepare for the development of an effective anti-hepatitis vaccine.

Methods

Tomato fruit-specific promoters' gene 2A12 and E8 were respectively introduced to pBPFΩ7 to form pB2A12 and pBE8. The DNA fragment containing HBsAg-s gene from plasmid YEP-HBs was inserted respectively into pB2A12 and pBE8 to form pB2A12-HBs and pBE8-HBs. The fragment containing "p35S+2A12+Ω+HBsAg-s+Tnos" of the pB2A12-HBs was sub-cloned into plasmid pCAMBIA1301 to yield the reconstructed plant binary expression plasmid pCAM2A12-HBs, and the fragment containing "p35S+E8+Ω+HBsAg-s+Tnos" of the pBE8-HBs was sub-cloned into plasmid pCAMBIA1301 to yield the plasmid pCAME8-HBs. The inserted gene HBsAg and fruit-specific promoters in the reconstructed plant binary expression vectors were confirmed by sequencing. Then, pCAM2A12-HBs and pCAME8-HBs were directly introduced into Agrobacterium tumefaciens strain EHA105.

Results

Digestion with restriction enzymes proved that all recombinant vectors had the inserts with expected length of the target fragments, and the sequencing results were confirmed correct.

Conclusion

In this study, plant expression vector containing HBsAg gene driven by fruit specific promoter and CaMV35s promoter was successfully constructed.

Key words: Promoter, 2A12, E8, Hepatitis B virus surface antigen, Plants

CLC Number: 

  • R512.6+2

Figure 1

Plasmid of plant expression vectors pCAME8-HBs and pCAM2A12-HBs A: pCAME8-HBs; B: pCAM2A12-HBs."

Figure 2

Restriction endonuclease analysis of pCAM2A12-HBs (PstⅠ) M: DNA Marker, Lamba DNA/Hind Ⅲ Markers; Lane 1, 2: pCAM2A12-HBs."

Figure 3

Restriction endonuclease analysis of pCAME8-HBs (PstⅠ)M: DNA Marker, Lamba DNA/Hind Ⅲ Markers; Lane 1, 2: pCAME8-HBs."

Figure 4

Restriction edonuclease analysis of plant binary vector pCAM2A12-HBs and pCAME8-HBs from Agrobacterium EHA105 (PstⅠ) Lane 1: DNA Marker, Lamba DNA/HindⅢ Markers;Lane 2~7: pCAM2A12-HBs; Lane 8~12: pCAME8-HBs."

Figure 5

PCR products of 2A12 and E8 promoters of plant binary vectors pCAM2A12-HBs and pCAME8-HBs from Agrobacterium EHA105 Lane 1: DNA marker, DL-2000; Lane 2: pCAMBIA1301; Lane 3, 4: pCAME8-HBs; Lane 5, 6: pCAM2A12-HBs."

Figure 6

PCR products of HBsAg fragments of plant binary vector pCAM2A12-HBs and pCAME8-HBs from Agrobacterium EHA105Lane 1: DNA marker, DL-2000; Lane 2: pCAMBIA1301; Lane 3, 4: pCAM2A12-HBs; Lane 5, 6: pCAME8-HBs."

[1] 毛自朝, 于秋菊, 甄伟 , 等. 果实专一性启动子驱动ipt基因在番茄中的表达及其对番茄果实发育的影响[J]. 科学通报, 2002,47(6):444-448
doi: 10.3321/j.issn:0023-074X.2002.06.012
[2] Pear JR, Ridge N, Rasmussen R , et al. Isolation and characterization of a fruit-specific cDNA and the corresponding genomic clone from tomato[J]. Plant Mol Biol, 1989,13(6):639-651
[3] Deikman J, Fischer RL . Interaction of a DNA binding factor with the 5'-flanking region of an ethylene-responsive fruit ripening gene from tomato[J]. EMBO J, 1988,7(11):3315-3320
[4] 王关林, 方宏筠 . 植物基因工程原理与技术(第1版)[M]. 北京: 科学出版社, 1998,( 第1版):547-549
[5] Vancanneyt G, Schmidt R , O'Connor-Sanchez A , et al. Construction of an intron-containing marker gene: splicing of the intron in transgenic plants and its use in monitoring early events in Agrobacterium-mediated plant transformation[J]. Mol Gen Genet, 1990,220(2):245-250
[6] Smigocki AC, Owens LD . Cytokinin gene fused with a strong promoter enhances shoot organogenesis and zeatin levels in transformed plant cells[J]. Proc Natl Acad Sci U S A, 1988,85(14):5131-5135
[7] Van Haaren MJ, Houck CM . Strong negative and positive regulatory elements contribute to the high-level fruit-specific expression of tomato 2A11 gene[J]. Plant Mol Biol, 1991,17(4):615-630
[8] Van Haaren MJ, Houck CM . A functional map of the fruit-specific promoter of the tomato 2A11 gene[J]. Plant Mol Biol, 1993,21(4):625-640
doi: 10.1007/BF00014546 pmid: 8448362
[9] Deikman J, Kline R, Fischer RL . Organization of ripening and ethylene regulatory regions in a fruit-specific promoter from tomato(Lycopersicon esculentum)[J]. Plant Physiol, 1992,100(4):2013-2017
[10] Good X, Kellogg JA, Wagoner W , et al. Reduced ethylene synjournal by transgenic tomatoes expressing S-adenosylmethionine hydrolase[J]. Plant Mol Biol, 1994,26(3):781-790
[11] Sandhu JS, Krasnyanski SF, Domier LL , et al. Oral immunization of mice with transgenic tomato fruit expressing respiratory syncytial virus-F protein induces a systemic immune response[J]. Transgenic Res, 2000,9(2):127-135
doi: 10.1023/A:1008979525909
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