Escherichia coli DH5α(pSCBE3-HF)|BNCC360499 |BNCC

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Escherichia coli DH5α(pSCBE3-HF)

Culture medium	LB + 50mcg/ml ampicillin: yeast extract 5.0g, peptone 10.0g,NaCl 10.0g, distilled water 1.0L,pH 7.0. Sterilization at 121 ℃ for 15min. After sterilization, ampicillin was added when the culture medium was cooled to 40-50 ℃ with a concentration of 50mcg/ml.
Subculture procedure	(1) Prepare a test tube containing 5~10mL of liquid medium and 2 plates; (2) Open it in the safety cabinet, heat the tip of ampoule in a flame, quickly drop sterile water to creak it, then break it with forceps; (3) draw 0.5mL of liquid culture medium into a freeze dried ampoule, fully rehydrate and transfer the solution to the liquid test tube, mix evenly; (4) inoculate a plate with 0.2mL of suspension liquid, repeat the step to obtain two plates; ⑤ Put all the liquid test tubes and plates under the above culture conditions for cultivation, and the strains can be used when they grow.
Growth conditions	37 ℃;18-24h; Aerobic
Storage conditions	2-8 ℃
Sharing mode	Public welfare sharing

Escherichia coli DH5α(pSCBE3-HF)

Storage conditions: 2~8 ℃

No. 360499

Product format: freeze dried,200ul

Validity period: 6 years

Biosafety level: 1,handle in ultra-clear table or safety cabinet

Receiving notice: if any abnormality is found on the day of receipt, please contact the customer service within 24 hours. If it is overdue, it is deemed that the product are well. The freeze dried culture shall be used up once and shall not be retained. Please operate in strict accordance with this instruction, otherwise the replacement of bacteria are not be available in case of aberrant growth and loss of viability.

Host bacteria:BNCC353719 DH5α

Plasmid:BNCC361031 pSCBE3-HF (contains kanamycin resistance gene).

Medium: LB resistant agar medium

Yeast extract 5.0g, peptone 10.0g, NaCl 10.0g, distilled water 1.0L, pH 7.0. Sterilize at 121℃ for 15min. After sterilization, add kanamycin at a concentration of 50mcg/ml when the medium is cooled to 40-50°C.

Growth conditions:37 ℃ aerobic 18-24h.

Recovery steps:

① Prepare 1 sterile liquid test tube (including 5~10mL of culture medium) and 2 pieces of 90mm plate;

② Open the ampoule in the biosafety cabinet, heat the tip of ampoule in a flame, quickly drop sterile water to creak it, then break it with forceps;

③ Draw 0.5ml of liquid medium into a freeze dried ampoule, fully dissolve and transfer the solution to the test tube of host bacteria solution , mix evenly;

④ Draw partial solution from the test tube and spread it over the plate at 200ul/plate;

⑤ Put all the test tubes and plates under the above specified conditions for cultivation. The plates are not allowed to seal by wrapping. After 18-24 hours, the culture medium is obvious turbid and the growth of plate colonies or moss occurs.

Plasmid purification:

1. Pick a single colony and inoculate it into 50ml liquid LB resistance medium, shake it for cultivation at 37 ℃ overnight;

2. According to the requirements, draw appropriate amount of bacterial liquid for plasmid extraction.