During our project, we designed, engineered, assembled, and tested many standard biological parts. Below you will find an introduction to our Best Composite Part and Best Part Collection candidates. You will also find the name, ID, and link to all the parts our team added to the registry.

Best Composite Part

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The SynLOCK Cassette (BBa_5087017) is a key component of SynLOCK—the crRNA Synthesis System for SHERLOCK, designed, built and tested by our team to simplify and standardize the process of generating custom crRNA molecules. The Cassette enables the creation of crRNAs with a 28-nucleotide spacer sequence tailored to the user's needs. This composite part makes our system a versatile and user-friendly platform for developing various detection tools that are compatible with simple Lateral Flow Assay tests and can be used directly in the field.

The SynLOCK Cassette is composed of several basic parts that together create a robust platform for crRNA synthesis:

• T7 promoter: Enables in vitro transcription using the plasmid carrying the SynLOCK Cassette as a template.

• Cas 13a DR (Direct Repeat): Allows the Cas13a protein to bind to the produced crRNA.

• SapI: Provides restriction sites for inserting a custom spacer sequence that is complementary to the detection target and excising the reporter for straightforward visual screening.

• Reporter unit: Positioned between the SapI cutting sites, the reporter unit enables differentiation between bacteria that did not incorporate the spacer sequence (reporter remains present) and those that successfully incorporated it (reporter is absent).

• BbsI: Offers a restriction site to linearize the plasmid carrying the SynLOCK Cassette, ensuring in vitro transcription of crRNA of the correct length.

Figure 1. Incorporation of a custom crRNA spacer into the SynLOCK Cassette.

Figure 2. Upon replacing the reporter with the custom crRNA spacer, colonies lose their purple color, making the screening process fast and straightforward.

Best Part Collection

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Our Best Part Collection is a carefully chosen set of parts that enable the creation of custom crRNA molecules for the SHERLOCK system. This collection was demonstrated to work in the detection of Prymnesium parvum.

With our platform, we wanted to address the time and cost inefficiencies associated with standard crRNA synthesis by ensuring that the process of crRNA design and generation is straightforward and accessible to future iGEM teams. SynLOCK simplifies the use of Cas13a proteins for scientists working with the SHERLOCK system, regardless of the target they want to detect. 

Parts of our collection come with two badges to make it easier to distinguish which part is a customizable element of the system, allowing the user to create their own crRNAs for various detection applications, and which is specific to Prymnesium parvum, serving as a demonstrative target to showcase the collection’s efficiency and applicability.

What You Can Find in the Collection

• Optimized set of RPA primers for detecting Prymnesium parvum (BBa_K5087000 - BBa_K5087005) and primers for the control assay (BBa_K5087010 and BBa_K5087011).

• CrRNA spacers for the SynLOCK system, allowing for the creation of crRNAs for the detection of Prymnesium parvum (BBa_K50019 and BBa_K50021) and a spacer for the control crRNA (BBa_K5087021).

• The SynLOCK Cassette (BBa_K5087017).

• Full crRNA molecule templates for the above-mentioned spacers to demonstrate their performance (BBa_K5087022 - BBa_K5087024).

• Two reporter modules—one that we used within our cassette (BBa_K5087025) and an alternative for further system customization (BBa_K5087026).

List of All Our Parts

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In this section, we present all the parts that our team added to the registry or contributed to on an existing part page. Our overall set includes:

• Primers designed and optimized by our team for RPA amplification of a genomic DNA sequence specific to Prymnesium parvum, as well as a few PCR primers for the same purpose.

• Building blocks for the easy construction of our SynLOCK system components, allowing for streamlined custom crRNA synthesis.

• crRNA templates that reflect the sequence of crRNA molecules needed to detect Prymnesium parvum, along with a positive control.

• Reliable chromoprotein reporters used in the SynLOCK system.

• A contribution to the LwaCas13a protein page.

Basic Parts

Primers

*PCR— Polymerase Chain Reaction, RPA — Recombinase Polymerase Amplification, IVT — In Vitro Transcription

SynLOCK System Building Blocks

CrRNA Templates

Composite parts

SynLOCK Cassette

Reporters

Contribution

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