[Construction of selectable marker-removable plant expression vectors].
The commonly used plant constitutive expression vector pBI121 was modified by insertion of two directly orientated lox sites each at one end of the selectable marker gene NPTII and by replacing the GUS gene with a sequence composed of multiple cloning sites (MCS). The resulting plant expression vector pBI121-lox-MCS is widely usable to accommodate various target genes through the MCS, and more importantly to allow the NPTII gene removed from transformed plants upon the action of the Cre recombinase. In addition, the CaMV 35S promoter located upstream of the MCS can be substituted with any other promoters to form plant vectors with expression features specified by the introduced promoters. Provided in this paper is an example that an enhanced phloem-specific promoter of the pumpkin PP2 gene (named dENP) was used to construct an NPTII-removable phloem-specific expression vector pBdENP-lox-MCS. Moreover, to facilitate screening of selectable marker-removed gene and the composite sequence is flanked by lox sites. Thus the selectable marker-free plants can be visually identified by loss of GFP fluorescence. The above newly created plant expression vectors can be used to develop selectable marker-removable transgenic plants for a variety of purposes.
Gao
Shang
S
State Key Laboratory of Plant Genomics, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100080, China.
Su
Lei
L
Jia
Hong-Ge
HG
Guo
Hong-Nian
HN
Tian
Ying-Chuan
YC
Fang
Rong-Xiang
RX
Chen
Xiao-Ying
XY
D001287:Attachment Sites, Microbiological / Q000235:genetics; D001665:Binding Sites / Q000235:genetics; D003001:Cloning, Molecular; D055786:Gene Knockout Techniques / Q000379:methods; D017343:Genes, Plant / Q000235:genetics; D005819:Genetic Markers / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D019426:Integrases / Q000235:genetics / Q000378:metabolism; D010944:Plants / Q000235:genetics*; D030821:Plants, Genetically Modified / Q000235:genetics*; D011995:Recombination, Genetic
D005819:Genetic Markers; D049452:Green Fluorescent Proteins; C045073:Cre recombinase; D019426:Integrases