Improving the electro-transformation efficiency of Corynebacterium glutamicum by weakening its cell wall and increasing the cytoplasmic membrane fluidity.
OBJECTIVE
To improve the transformation efficiency of Corynebacterium glutamicum cells with heterogenous plasmid DNA and single-strand DNA (ssDNA) using a methodology based on electro-transformation.


RESULTS
A semicomplex hypertonic medium was selected with addition of glycine and DL-threonine to weaken cell walls and addition of Tween 80 and isonicotinic acid hydrazide to increase cytoplasmic membrane fluidity. Their contents were optimized by response surface methodology. Cell growth, electro-transformation buffer, and transformation protocol were also optimized. Temporary heating inactivation of the host restriction enzyme showed a significant effect. Finally, a high transformation efficiency of 3.57 ± 0.13 × 10(7) cfu/μg DNA of plasmid and 1.05 × 10(6) Str (R) cfu per 10(9) viable cells with a ssDNA was achieved.


CONCLUSIONS
The results shed light on the application in functional genomics and genome editing of C. glutamicum.
The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, No 1800 Lihu Avenue, Wuxi, 214122, Jiangsu, China. summer.ruan19@gmail.com.
Yili
Y
Ruan
The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, No 1800 Lihu Avenue, Wuxi, 214122, Jiangsu, China. zlj@jiangnan.edu.cn.
Linjiang
L
Zhu
The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, No 1800 Lihu Avenue, Wuxi, 214122, Jiangsu, China. liqi@jiangnan.edu.cn.|State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi, 214000, China. liqi@jiangnan.edu.cn.
Qi
Q
Li
D002462:Cell Membrane / Q000378:metabolism; D002473:Cell Wall / Q000378:metabolism; D048230:Corynebacterium glutamicum / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D003470:Culture Media / Q000737:chemistry; D018274:Electroporation / Q000379:methods*; D007538:Isoniazid / Q000378:metabolism; D008560:Membrane Fluidity / Q000187:drug effects; D010957:Plasmids; D011136:Polysorbates / Q000378:metabolism; D014169:Transformation, Bacterial
D003470:Culture Media; D011136:Polysorbates; D007538:Isoniazid
Cell-wall weakening; Corynebacterium glutamicum; Cytoplasmic membrane fluidity; Electro-transformation; Single-strand DNA