Product Specification
MC1061F-：100μl/tube
pUC19 (control vector，10pg/μl): 10μl
storage condition (Guarantee period): -80℃（6 months）
Genotype
araD139 Δ (araA-leu)7697 Δ(lac)X74 galK16 galE15(GalS) lambda- e14- mcrA0 relA1 rpsL150
(StrR) spoT1 mcrB1 hsdR2
Product Description
The MC1061F- strain was derived from E. coli B/r SB3118 strain, and was also the parent strain of DH10b and TOP10. MC1061F - high conversion efficiency of transfer cells, and can be used for conventional plasmid of laboratory building and phage display experiment, but the lack of F ` factor, can not be used in filamentous phage (such as M13) infection. EndA1 mutation does not contain nuclease, and the content of nuclease in vivo is high. When extracting plasmid, it is important to use the albumin removal solution in plasmid extraction kit to remove the contamination of nuclease to plasmid as much as possible. This strain is resistant to streptomycin. The MC1061F- sensitized cells produced by vivo were made by a special process. The conversion efficiency of pUC19 plasmid detection and transformation was > 2*109 cfu/μg DNA.
Operation Method
1. MC1061F - cells from -80 ℃, inserted rapidly in the ice, 5 minutes to the fungus block melt, join the target DNA (plasmid or connection product) and hand dial EP pipe bottom gently blending (avoid with a suction gun), ice stand for 25 minutes.
2. 42 ℃ water bath heat shock 45 seconds, quickly back to the ice and let stand for 2 minutes, shaking will lower conversion efficiency.
3. Add 700 μl of sterile medium (2YT or LB) without antibiotic to the centrifuge tube, after blending, place it in the shaker at 37 ℃, 200 RPM for 60 minutes cell recovery.
4. The bacteria were collected by centrifugation at 5000 RPM for 1 minute. In the centrifuge tube, left aproxiamatly 100 μl of supernatant, which was used to gently blow the heavy suspensions and smear them on the 2YT or LB medium containing the corresponding antibiotics.
5. Put on the plate up side down in the 37 ℃ culture box over night. For the blue white spot filtering operation, place the culture plate in 37 ℃ for at least 15 h.
Matters needing attention
1.The receptor cells are best melted slowly in ice. Insert the target DNA within 8 minutes on ice. Do not put it in the ice for too long. Long-term storage will reduce the conversion efficiency.
2. The conversion of high concentration of plasmids or high-efficiency bonding products can reduce the amount of bacteria eventually used for coating.
