HARDWARE

During the project's advancement, we recognised that deploying the product in practical applications urgently required developing a field-operable detection tool capable of rapidly assessing salicylic acid levels in soil. Existing mainstream techniques predominantly rely on laboratory analysis, involving multiple steps such as sampling and submission for testing, which fails to meet the requirements for real-time, in-situ detection. Given that salicylic acid serves as the key inducer regulating the secretion of fusion proteins by engineered bacteria within this system, its concentration level directly determines the successful secretion of the target protein. Consequently, establishing an efficient and convenient method for salicylic acid detection holds significant importance for the project's implementation.

Accordingly, we have designed a device specifically for rapid salicylic acid detection in soil. The device consists of two main components: an external protective sampling sleeve and an internal detection tube containing salicylic acid-specific test strips. During operation, the device is first fully inserted into the target soil area. Water is then injected through the sleeve in appropriate quantities to ensure thorough contact with the surrounding soil, forming a soil leachate. The detection tube is subsequently placed into the sleeve. Under pressure, the leachate contacts the detection paper strip and triggers a reaction. The resulting color change enables a semi-quantitative assessment of salicylic acid content in the soil, providing critical data support for subsequent engineering applications.

The sleeve, as the core component of this detection apparatus that comes into direct contact with the soil, primarily serves to provide structural protection and operational guidance for the internal detection tube and salicylic acid test paper. This component features an elongated structure with an external diameter of 20 mm and a total length of 250 mm. This design facilitates smooth penetration of the apparatus into diverse soil conditions, minimising operational resistance.

When determining the sleeve dimensions, we comprehensively considered the root distribution characteristics of leguminous plants alongside the material's mechanical properties. Available data indicates that the primary root zone of most leguminous plants typically extends to a depth of 600 mm or less. Concurrently, based on the mechanical strength characteristics of PLA (polylactic acid) material, setting the sleeve length to 250 mm effectively covers the detection requirements for the typical root zone of leguminous plants. This length also complies with ergonomic principles, facilitating field insertion and removal operations for users.

To effectively constrain the axial movement of the test tube within the sleeve, a countersunk hole structure is incorporated internally. This facilitates precise positioning and mechanical limitation of the test tube. This design prevents excessive downward displacement of the test tube during operation, thereby avoiding damage to the salicylic acid test paper and the test tube itself due to pressure or impact. The countersunk hole structure features a wall thickness of 2 mm at the upper section and 4 mm at the lower section, ensuring overall structural integrity while optimising material distribution. Furthermore, the bottom wall of the sleeve is tapered to form a conical insertion end, significantly reducing resistance during soil penetration. This facilitates smoother operation and enhances penetration efficiency.

To facilitate the replacement of test strips and enhance the overall maintainability of the structure, the test tube employs a modular design concept, dividing it into four functionally independent components, as illustrated in the figure. From top to bottom, the four structures are designated as 1 (Fig. 4), 2 (Fig. 5), 3 (Fig. 6), and 4 (Fig. 6). (Structures 3 and 4 are identical.)

The top component of the test tube is a cylindrical structure with an external diameter of 16 mm. This dimension, determined through ergonomic considerations, facilitates secure grasping and force application by the operator. Furthermore, this external diameter forms a mating relationship with the 16 mm internal diameter at the upper end of the countersunk hole within the sleeve. This provides effective axial positioning of the test tube, preventing excessive downward displacement and protecting the test strip and bottom filter membrane from mechanical damage.

Salicylic acid test paper is fixed between the upper end of component 1 and the middle section of component 2, ensuring its stable positioning during testing. The middle section employs a stepped shaft design: its upper half maintains a 16 mm outer diameter consistent with the top component, while the lower half features a 12 mm outer diameter, facilitating a smooth structural transition. The middle component connects to two identical tail components via three interfaces. Between these interfaces, a semi-permeable membrane, a layer of standard filter paper, and a metal mesh are sequentially interposed, collectively forming a multi-stage filtration system. This enhances the selectivity and uniformity of liquid permeation during testing. The total length of the test tube is 275 mm, balancing detection depth with operational convenience.

To connect components, a specialised locking mechanism is incorporated at the tube head (Fig. 7), with corresponding assembly space reserved at the tube tail for rapid engagement. All locking mechanisms employ uniform dimensions and geometry, enhancing assembly consistency while facilitating future functional expansions or structural improvements. This design enhances the device's reconfigurability and long-term utility.

First, position the salicylic acid detection paper strip onto the snap-fit structure at the upper end of Component 2. Subsequently, connect Component 1 to Component 2 via the snap-fit mechanism, securing the detection paper strip between the joint interfaces of both components. Employing the same assembly method, place the semi-permeable membrane between Components 2 and 3, completing the connection via the snap-fit. Finally, insert standard filter paper between Components 3 and 4, securing it using the same method. Position the metal mesh at the base of Tube 4 to complete the assembly of the full Test Tube II.

After identifying the test area, insert the sharpened end of Tube I (Sleeve) into the soil, ensuring the tube protrudes no more than 5 cm above ground level. Pour an appropriate volume of clean water through the upper opening of Tube I until the soil within is fully saturated. Subsequently, insert the fully assembled Tube II axially along Tube I, applying moderate pressure to facilitate the upward migration of leachate exuded from the saturated soil under compression. Maintain pressure until the liquid surface contacts the test paper strip. After allowing the reaction to proceed, assess the approximate salicylic acid concentration in the soil based on the resulting colour change of the test paper.

During the preliminary research and conceptual design phase, we focused on developing a portable device for efficient detection of salicylic acid content in soil, while exploring its functional expansion potential. This aimed to enable rapid detection of multiple target substances by swapping different types of test strips. Balancing portability, structural rationality, and operational convenience, we initially established a cylindrical configuration for the device's overall structure./

The initial design specified an outer diameter of approximately 50 mm and a total length of around 180 mm. To validate this design, a preliminary physical prototype was produced using 3D printing technology. During practical simulation testing, this prototype revealed two primary issues: Firstly, to accommodate printing simplicity, the sleeve's inner wall thickness was uniformly set at 5 mm. This resulted in an overall structure that was disproportionately short and thick, leading to significant insertion resistance when driven into soil. It was easily obstructed by surface gravel and plant root systems. Secondly, the sleeve's substantial external diameter risked causing significant disturbance or mechanical damage to existing root systems when deployed near legume root zones, potentially compromising the representativeness of detection results and adversely affecting plant growth./

To address these concerns, we engaged in thorough discussions with Professors Chunlei and Peng Huan. Under their guidance, we implemented crucial structural optimisations: reducing the outer diameter to 20 mm while increasing the sleeve length to 250 mm. This adjustment significantly enhances penetration capability in complex soil conditions while minimising potential interference with plant root systems./

Subsequent field use revealed that inserting the detection tube into the sleeve could inadvertently cause excessive penetration due to awkward force application, posing a risk of damaging internal test strips. To address this, we incorporated countersunk holes into the sleeve's inner wall. A 2 mm height differential between the upper and lower inner surfaces provides mechanical limitation for the detection tube. Following multiple rounds of field validation, this dimensional design effectively controls the detection tube's travel while ensuring the sleeve maintains sufficient mechanical strength and durability. This guarantees reliable operation of the device in field conditions./

In the initial design proposal, to enhance the convenience of water injection operations, we planned to integrate a three-way water injection structure (referred to as the ‘three-way pipe’) on the right side of the sleeve to enable portable water replenishment. However, subsequent structural refinement and mechanical analysis revealed that to ensure water flow could be effectively directed into the underlying soil, the injection pipe required a relatively small internal diameter. Yet, an excessively narrow pipe profile would significantly compromise its mechanical strength, making it susceptible to fracture under lateral forces during the sleeve's penetration into the soil./

To resolve this conflict, we attempted to enhance structural stability by increasing wall thickness or optimising the cross-sectional shape, designing several improved iterations accordingly. However, these modifications invariably increased structural complexity to some degree, while still struggling to simultaneously satisfy the dual requirements of fluid permeability and mechanical reliability within the limited space./

Following in-depth discussions and comprehensive evaluation with Professor Chunlei, we concluded that the three-way injection structure not only elevates manufacturing costs and process complexity but also poses potential risks to operational reliability in field conditions. Consequently, the final design omits this component, adopting a simpler and more dependable top-end direct injection method for water replenishment. This approach ensures functional fulfilment while optimising the overall structure's robustness and practicality./

During the design phase of the locking connector structure for the detection tube, we proposed two viable technical solutions and conducted a systematic evaluation:/

Solution One: External Snap-Fit Connection. This design employs a pair of symmetrically distributed cantilevered snap-fit structures, achieving rapid assembly by engaging with pre-set corresponding notches in the tube wall. This solution offers the advantages of a secure connection and high structural stability, effectively withstanding axial loads to enhance the device's durability and service life. Its drawback lies in the interference fit between the clips and notches, requiring substantial external force for disassembly, which slightly compromises maintainability./

Option Two: Internal Slot Connection. This design incorporates a thin-walled annular flange at the tube end, with a matching annular groove pre-machined into the inner wall of the corresponding connector. Securing is achieved through axial press-fitting. The primary advantage of this structure lies in the complete concealment of the connection within the pipe, offering superior visual integrity, concealment, and aesthetics. However, due to the inherent mechanical properties of PLA material, the thin-walled flange structure formed by printing is prone to brittle fracture under stress, leading to connection failure. This severely compromises the reliability of this solution in practical applications./

Upon comprehensive comparison of the engineering applicability of both approaches, Solution Two exhibits significant structural strength deficiencies, rendering it unsuitable for meeting durability requirements in field inspection environments. Consequently, Solution One was ultimately selected as the implementation method for the locking mechanism. This solution, with its superior connection strength and structural robustness, better aligns with the core requirements for hardware reliability and long-term stability demanded by this apparatus./

To address potential process precision limitations in 3D printing for the latch structure under Option One, we have concurrently developed a preparatory manufacturing approach: designing and printing the latch structure and the main body of the inspection tube as separate components, followed by post-assembly to achieve a reliable connection between the two. This alternative approach effectively mitigates the risk of moulding defects arising from overhanging structures or dimensional deviations inherent in monolithic printing. While ensuring structural functional integrity, it significantly enhances manufacturing success rates and process tolerance, providing reliable technical redundancy for the project's continued advancement./

During the hardware development process, we first completed preliminary structural design sketches through multiple rounds of internal team discussions. Subsequently, we engaged in in-depth exchanges with Professor Liu Jianmiao regarding the initial draft. She proposed designing the detection tube as a segmented structure, a concept that effectively expanded the device's modular potential, enabling adaptation to more diverse application scenarios. Guided by this directional feedback, we systematically refined the hardware design./

To refine manufacturing process details, we specifically consulted faculty members specialising in 3D printing technology at the Engineering Practice Innovation Centre of Huazhong University of Science and Technology. We conducted specialised discussions focusing on print precision, material properties, and structural feasibility. Following their professional advice, we made precise adjustments to the dimensional parameters of the locking mechanism and the inclination angles at connection points. This significantly enhanced assembly reliability and structural longevity. After multiple rounds of iterative optimisation and comprehensive evaluation, we finalised a hardware design solution with high engineering feasibility./

Below are additional drafts and sketches from our hardware design process. Images are hosted on tools.igem.org; the links below point to the uploaded files in your gallery.

There are also several PDF files in the same gallery. You can download them here:

• drafts-in-hardware-design-2.pdf (Preview / download)
• drafts-in-hardware-design-3.pdf (Preview / download)
• drafts-in-hardware-design-4.pdf (Preview / download)
• drafts-in-hardware-design-5.pdf (Preview / download)
• drafts-in-hardware-design.pdf (Preview / download)