Experimental Samples

Source of experimental samples

Strains and carriers used in the experiment

Strains and plasmids used

Main chemical reagents in the experiment

Reagent formulations

(1) Luria-Bertani ((LB) medium: peptone 10g / L, yeast extract 5g / L, NaCl / 10g / L, pH 7.0121 ℃, 20 min), solid LB medium plus 2% (Wamp V) Agar.

(2) Inorganic salt culture medium (MSM) with polyethylene as the sole carbon source: KH2PO4 0.7g shock L, K2HPO4 0.7g shock L, MgSO4 ·7H2O 0.7g K2HPO4 1.0g shock L, FeSO4 ·7H2O 0.002 g shock L, ZnSO4 ·7H2O 0.002 g max L, MnSO4 ·H2O 0.01g max L. pH 7.0 After sterilization at 121℃for 20 min, polyethylene (PE) film with 3 × 3 cm and PE microsphere with a diameter of 30 μ m were added for 10 mg respectively. Pretreatment of the film and polyethylene microspheres: 90% ethanol (1 h) and 75% ethanol (1 h) treated the film (3 cm × 3 cm) and 30 μ m microsphere particles and dried before use (40 °C, 30 min). The treated polyethylene samples were cultured in LB (pH 7. 0) for 24 h to confirm that they were fully sterilized.

(3)50 × TAE buffer: weighing 242gTris-base,37.2 g Na2EDTA ·2H2O, then adding 800 mL distilled water, fully stirring and dissolving, then adding 57.1 mL acetic acid, fully mixing, finally distilled water fixed volume to 1 L, room temperature. Dilute it to 1×TAE when in use (for example, if you need to use 1 × TAE of 1000 mL, then take 50 × TAE,980 mL of deionized water of 20 mL, mix and rest for a while before use).

(4) IPTG storage solution: weigh 2 g isopropylthiogalactoside solid, dissolve it in 8 mL distilled water, configure it into 50 mg/mL storage solution, use 0.22 μ m filter membrane to remove bacteria in a 2 mL centrifuge tube and store it in a refrigerator at-20 ℃ for use.

(5) preparation of Kana solution: 0.5g kanamycin sulfate solid was dissolved in 9 ml distilled water and configured as 50 mg/mL storage solution. 0.22 um filter membrane was used to remove bacteria in 2 mL centrifuge tube in super clean table, and stored in refrigerator at-20 ℃. Its working concentration was 50 μ g/mL.

(6) Coomassie brilliant blue dye: 1 g Coomassie brilliant blue R250 was dissolved in 400 mL ddH2O, 450 mL methanol, 100 mL glacial acetic acid was added, and finally ddH2O was added to fix the volume to 1 L.

(7) SDS-PAGE decolorizing solution: methanol 100mL, glacial acetic acid 100mL, ddH2O volume 1L.

(8) SDS-PAGE protein loading buffer (5 × SDS-PAGE Protein Loading Buffer): 1.25 mL 0.5 mol/L Tris-HCl pH6.8, glycerol 2 mL, 10%SDS 2 mL β-mercaptoethanol 1 mL, 0.1% bromophenol blue 0.5 mL, dissolved in distilled water to 10 mL, then packed the prepared solution into 1.5 mL EP tubes, 1 mL per tube, stored in a refrigerator at-20 ℃.

Bacterial colony PCR

PCR Reaction system

PCR Amplification reaction conditions