Making it Rain

Playing RPS with E. coli during summer was fun, but, even if humans won, celebrations did not last long since we soon returned to complaining about the hot weather. As a prize for humans who win in our RPS game, we designed an E. coli that can make it rain, making the hot summer more fun and refreshing (let alone applications in agriculture).

1. Introduction

To make it rain we focus on the substance isoprene. It has been observed that trees in tropical rainforests contribute to the formation of photo-smog aerosol in the lower atmosphere by releasing isoprene (Paulson and Seinfeld, 1992). The photo-oxidized isoprene acts as a condensation nucleus [2], may cause shower (make it rain) even if it is present in very low concentrations.

Fig1. Isoprene photo-oxidation reaction

It is known that the enzyme isoprene synthase can catalyze the conversion of dimethylallyl diphosphate(DMAPP) to isoprene. DMAPP is normally synthesized by E. coli, so the only thing we need to make our bacteria synthetize isoprene is isoprene synthase. The isoprene synthase coding gene (ispS) is isolated from the tree poplar (Barbara Miller et al., 2001). E. coli introduced this gene released isoprene into the air by diffusion [1]

Fig2. Formation of isoprene is catalyzed by isoprene synthase

In this study, we tried to make E. coli synthesize isoprene by introducing ispS.

2. Isoprene by E. coli

To measure the amount of isoprene produced by E. coli introduced ispS, we constructed negative control PlacIQ and sample PlacIQ-rbs-ispS, using the PlacIQ promoter (BBa_I14032) and ispS. Gene ispS is extracted from the pMK backbone vector. (see more about our constructions)

Fig3. Constructions of rbs-ispS and PlacIQ-rbs-ispS

We used Gas Chromotrography-Mass Spectrometry (GC-MS) to measure the amount of isoprene from E. coli. When using GC-MS, we injected a series of chloroform-diluted liquid isoprene to draw the calibration curve. To confirm that liquid isoprene produced by E. coli would be released as a gas, we diluted liquid isoprene in water and also in LB medium. In both cases, we could confirm isoprene was evaporated into the air (see more about these experiments).

According to the calibration curve, we detected 4.1×10^-5 mg/L isoprene produced by E. coli BL21 (DE3) introduced isoprene synthase, while negative control (PlacIQ) produced one eighth of our new E. coli. (see more about GC-MS)

(a) (b)

3.Discussion

The reaction between isoprene and ozone has been studied to examine physical and chemical characteristics of the secondary organic aerosol formed. Aerosols is suspension of solid particles or liquid droplets in gas. The most common aerosols in the atmosphere are clouds, which normally consist of suspensions of water droplets or ice particles of greater density, and can later cause rain. According to those information, we designed an easy indoor experiment of reaction between isoprene and ozone, and confirmed that isoprene can make aerosol. (see more...) All in all, we confirmed that E. coli with the insertion of ispS synthesizes isoprene and that isoprene makes aerosol. So our E. coli will make it rain!