Construction of a plant expression vector of chalcone synthase gene of <italic>Ginkgo biloba</italic> L. and its genetic transformation into tobacco

doi:10.1007/s11703-010-1033-7

Front Agric Chin

2010, Vol. 4

Issue (4) : 456-462 https://doi.org/10.1007/s11703-010-1033-7

RESEARCH ARTICLE

Construction of a plant expression vector of chalcone synthase gene of Ginkgo biloba L. and its genetic transformation into tobacco

Linling LI^1,2, Hua CHENG², Jianying PENG¹(

), Shuiyuan CHENG²(

)

1. College of Horticulture, Agricultural University of Hebei, Baoding 071001, China; 2. College of Life Science and Engineering, Huanggang Normal University, Huanggang 438000, China

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Abstract

The chalcone synthase gene (chs) from Ginkgo biloba L. was cloned by PCR procedure. For constructing a plant expression vector of Gbchs, the gene was digested with XbaI and BamHI and inserted into the pBI121 vector. Gbchs was transferred into tobacco mediated by Agrobacterium tumefaciens LBA4404. PCR and Southern blot were performed, and the results showed that chs had been transformed into the genomic DNA of tobacco. The total flavones in the transformed tobacco leaves was extracted by the methanol extraction method and tested by UV spectrophotometry. The results showed that, compared to the control tobacco, the content in the transgenic tobaccos, n=6, was generally higher, and the difference reached a significant level (P<0.05) and highly significant level (P<0.01), except one sample. Moreover, the highest sample was nearly 7.7 times greater than that of the controls. All these results show that utilizing genetic manipulations to improve plants in order to regulate flavone content by gene engineering may be an effective and hopeful method.

Keywords chalcone synthase gene Ginkgo biloba L. plant expression vector flavones

Corresponding Author(s): PENG Jianying,Email:pengzhuwen@sina.com; CHENG Shuiyuan,Email:s_y_cheng@sina.com

Issue Date: 05 December 2010

Cite this article:

Linling LI,Hua CHENG,Jianying PENG, et al. Construction of a plant expression vector of chalcone synthase gene of Ginkgo biloba L. and its genetic transformation into tobacco[J]. Front Agric Chin, 2010, 4(4): 456-462.

URL:

https://academic.hep.com.cn/fag/EN/10.1007/s11703-010-1033-7
https://academic.hep.com.cn/fag/EN/Y2010/V4/I4/456

Fig.1 Colony PCR detection of pBI121-
Note: 1-5: PCR product of positive colonial; CK-: PCR product of wild colonial; M: DL2000 marker.

Fig.2 The restriction enzyme digestion analysis of pBI121-
Note: M: DL2000 marker; 1: the restriction enzyme digestion of recombinant vector; 2: .

Fig.3 Colony PCR detection of pBI121- in LBA4404
Note: + : PCR product of pBI121; -: PCR product of wild LBA4404 colonial; 1-4: PCR product of positive colonial; M: DL2000 marker.

Fig.4 Identification of transgenic tobaccos by PCR
Note: + : PCR product of pBI121-; -: PCR product of control tobacco; M: DL2000 marker; 1-8: PCR product of Kanamycin-resistant plants.

Fig.5 Southern blot analysis on putative primary transformants of tobaccos
Note: CK: control tobacco; 1-5: positive tobaccos by PCR analysis.

Fig.6 Transcript accumulation of the in transgenic or control tobaccos by RT-PCR (upper panel)
Note: The gene was used as the control to show the normalization of the amount of templates in PCR reactions (lower panel). CK: control tobacco; sample 1-6: transgenic tobaccos.

Fig.7 The content of flavone in the transgenic or control tobacco leaf in net weight/%
Note: CK: control tobacco; sample 1-6: transgenic tobaccos. The difference reached significant level (<0.05) marked with *, very significant level (<0.01) marked with **.

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