Preparation method of xanthomonas campestris competent cells
A technology of competent cells and Xanthomonas, applied in the field of genetic engineering, can solve the problems of inability to transform and low transformation efficiency, and achieve the effect of good condition, high transformation efficiency and improved transformation rate
Inactive Publication Date: 2020-08-14
INNER MONGOLIA UNIV OF TECH
View PDF5 Cites 0 Cited by
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Since this method is mainly aimed at Escherichia coli, when it is applied to Xanthomonas campestris cells, there is a problem that the transformation efficiency is low or even cannot be transformed.
Method used
the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View moreExamples
Experimental program
Comparison scheme
Effect test
Embodiment Construction
[0025] The present invention will be described in detail in conjunction with the following specific examples. The following examples are not limitations of the present invention, and the protection scope of the present invention cannot be limited thereby.
[0026] The invention relates to a preparation method of Xanthomonas campestris competent cells with simple operation, which mainly includes the following steps:
[0027] 1) Pick a single colony of activated Xanthomonas campestris from the plate and put it into 10mL YYRZ liquid medium, culture it with shaking at 220rpm at 30°C for 12-16h;
[0028] 2) Transfer 1 mL of the overnight cultured bacterial solution to a 50 mL Erlenmeyer flask containing 10 mL of YYRZ liquid medium, and culture at 30°C with shaking at 220 rpm until the OD600 value is 0.4 to 0.6;
[0029] 3) Divide 10mL of the bacterial liquid into two 10mL pre-cooled sterile centrifuge tubes, centrifuge at 4000rpm for 10min at 4°C, discard the supernatant, absorb th...
the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More PUM
Login to View More Abstract
The invention discloses a simply-operated preparation method of xanthomonas campestris competent cells. The preparation method comprises the following steps of selecting a single xanthomonas campestris single colony from a flat plate, inoculating the single xanthomonas campestris single colony into 10mL of YYRZ liquid culture medium, and carrying out shake culture at 30 DEG C and 220rpm overnight;transferring 1mL of bacterial liquid cultured overnight into 10mL of YYRZ liquid culture medium, performing shake culture at 30 DEG C and 220rpm until an OD600 value is 0.4-0.6, standing the obtainedbacterial liquid on ice for 30min, centrifuging at 4 DEG C, discarding supernatant, and recovering thalli; adding a pre-cooled LHGG solution, re-suspending the thalli, carrying out ice bath for 30 minutes, centrifuging at 4 DEG C, discarding supernatant, recovering the thalli, and repeating the step once; and then adding a precooled LHGG solution to resuspend the thalli, subpackaging the thalli in 1.5 mL EP tubes with a volume of 200 [mu] l / tube, and preserving the thalli at a temperature of -70 DEG C for later use. The method is simple to operate, high in conversion efficiency and suitable for P1-grade microorganism laboratories.
Description
technical field [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to a preparation method of Xanthomonas campestris competent cells with simple operation. Background technique [0002] The competence of Xanthomonas campestris refers to a physiological state in which Xanthomonas campestris is processed to become receptive to exogenous DNA fragments. Untreated Xanthomonas campestris cells cannot absorb exogenous DNA. Therefore, the preparation of competent cells is an important link in genetic engineering operations, which directly affects the subsequent experimental work. At present, the competent cells prepared by conventional methods are Escherichia coli as the object, and chemical reagents such as CaCl2 and RuCl are used to expand the cell wall of Escherichia coli, and the permeability of the cell membrane is changed, so that foreign DNA can enter. Such cells are competent cells. (competent cell). Since this method is m...
Claims
the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More Application Information
Patent Timeline
Login to View More Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N1/38C12R1/64
CPCC12N1/20C12N1/38
Inventor 薛艳军李文彬刘丽华
Owner INNER MONGOLIA UNIV OF TECH