Building Blocks: Large DNA/Genome assembly made easy!

Overview

The Johns Hopkins University Build a Genome team has proposed a new standard, RFC38, for the construction and assembly of large DNA. The physical assembly of standard parts is currently a non-standard process, which can either come from direct genome PCR with restriction enzyme sites incorporated into the PCR primers, or overlap assembly PCR. Furthermore, current Biobrick construction standards [RFC10, RFC11,RFC12 etc.] rely heavily on restriction enzyme based methods, which can be sequence, cost and time restrictive. This is especially true of large DNA assembly, where 100% control of over DNA sequence may be mandatory, such as in whole genome assembly. We propose a novel standard, The Building Block Method, for both the construction of standard parts and their assembly

This Building Block standard can be either:

• Interchangeable
• Non-interchangeable format, based on the desired use of the part.

What is a Building Block?

• A Standard Build Block is a ~750 base pair sequence of DNA that is composed of annotated biological parts. This particular size represents a “sweet spot” for a currently optimized set of efficient assembly techniques (e.g. high accuracy sequencing in a single run) but need not be fixed in stone. The Building Block, however, may not be composed entirely of single biological parts (as in BioBricks) and the junction with neighboring Building Blocks may be essential for biological for function.

• A Composite Building Block is composed of two or more Building Blocks. Composite Building Blocks may be constructed from non-interchangeable Building Blocks or interchangeable Building Blocks.

Interchangability

• An Interchangeable Building Block contains standard sequences on both 3’ and 5’ends

• A non-interchangeable Building Block MAY contain non-standard 3’ or 5’ ends to aid in the construction of Composite Building Blocks.