Cloning of SjCA gene and its expression analysis on upland cottons

Abstract

Acquisition of salt-tolerant genes from exogenous plants to improve cotton salt resistance has always been a hotspot of research on cotton salt resistance. However, the information regarding the method of conversion of living cotton pollens by portable gene gun technology is still little. Complete sequence information of the Carbonic Anhydrase (CA) gene was obtained from NCBI database, and its full ORF length sequence was cloned by RT-PCR technology. After the construction of pBI121-CA::GFP fusion expression vector, we used cotton pollens from upland cotton varieties Y-2067, ZA-23, and GZ-2, which have weaker autofluorescence, to conduct the research on the transient expression of cotton pollens in vivo via particle bombardment (gene gun technology). The results indicated that green fluorescence enhancement of the three kinds of cotton pollens was realized after the CA gene transformation, meaning that the CA gene expression level was increased. Besides, the salt tolerance germination ability of transgenic T1 seeds was also improved. Our research initially established a transient expression system of cotton pollen in vivo via particle bombardment technology, laying the theoretical foundation for further research of cotton genetic transformation and creation of cotton salt-tolerant germplasm.

Cloning and Ectopic Expression of <i>ScYCF</i>1 Gene from <i>Saccharomyces cerevisiae</i> in Cotton

“…The powders and plasmids were treated as the way of Kong Jingjing [14] and the particle gun transformation in vivo was completed.…”

“…The germination experiment was performed on obtained T 0 seeds with the double walled filter paper method [14], using 100 mM NaCl solution as treatment solution and water as control. Each group repeated for three times.…”

“…Under excitation light at a wavelength of 488 nm, the auto-fluorescence of cotton pollen is green [14]. According to previous findings, three varieties Y2067, ZA-23 and GZ-2 with weak auto-fluorescence [14] were selected to collect pollen, and gene gun in vivo conversion technology was used to bombard the cotton pollen.…”

“…Under excitation light at a wavelength of 488 nm, the auto-fluorescence of cotton pollen is green [14]. According to previous findings, three varieties Y2067, ZA-23 and GZ-2 with weak auto-fluorescence [14] were selected to collect pollen, and gene gun in vivo conversion technology was used to bombard the cotton pollen. The study found that after the vector pBI121-ScYCF1:GFP was bombarded, the green fluorescence of pollens of three materials was significantly enhanced (Figure 6), indicating that ScYCF1 can be expressed in cotton pollen, which lay the foundation of using gene gun in vivo conversion technology to obtain transgenic salt-tolerant materials in the next step.…”