Table 1.

Supported functions (Operations/nodes). These functions are available as buttons on the main menu bar at the top of the GUI.

Function (Node)	Comments
DNA sequence	Load DNA sequence files into MCDS flowchart to create a new node
Restriction digestion	Use DNA in selected node(s) as substrate(s) for restriction digestion. If sites of two or more enzymes overlap, MCDS will warn user about the confliction
PCR	Use DNA in selected node(s) as template to perform PCR
	MCDS will generate single strand intermediates and then calculating the double strand products by annealing the intermediates. Therefore, MCDS generates by-products to allow users to understand the performance of their primers.
Modification	Apply CIAP (calf intestinal alkaline phosphatase)/Klenow Fragment blunting/T4 DNA polymerase blunting/polynucleotide kinase to selected nodes
Gel electrophoresis	Simulate gel electrophoresis for separation of DNA fragments
Ligation	Apply ligation algorithm to DNA from all selected nodes to generate ligation products
Screening	Use feature screening (simulates e.g. antibiotic resistance) or PCR screening to pick DNA of interest from a mixture
	Feature screening: MCDS detects all features in the substrates and list them in the property panel. Users can specify a combination of features and number of appearance. MCDS will work out the DNAs that match the conditions
	PCR screening: MCDS will use each of substrate to produce PCR products with given primers. Substrates that can generate PCR products within specific length range will be selected
Recombination	Apply recombination algorithm to DNA from all selected nodes to generate recombination products
Restriction enzyme analysis	Apply restriction enzyme analysis to DNA form all selected nodes. Users can set up conditions
Sequence merging	Use merging algorithm to merge two sequences with overlapping ends (e.g., overlapping sequencing results). Generates all possible outcomes if multiple homologous sequences are present
Sequence designer	Create a node that allows the user to design any DNA sequence by typing, pasting, or using a feature
	The designer code syntax can be found in the table online: https://goo.gl/qNdZa0. There is also an alternative UI sequence designer
Sequence information hash code selection	This function assigns a ‘hash code’ to all sequence products of a function (e.g. ligation, recombination). It allows the user select desired sequences where length or features (e.g. selectable markers) cannot be used to differentiate between target fragments
Sequencing simulation	Create a node that can generate the theoretical sequence of a one-primer sequencing reaction
Sequence comparison	Create a node that compares DNA from selected nodes. In the sequence comparison node, users can select one sequence as template and other sequences will be aligned to the selected sequence. Both features and comparison alignments are be shown in the sequence viewer and vector map to allow users to check if an alignment (i.e. sequencing result) covers a feature (i.e. gene)
Host	Create a node which define a host cell. Useful to confirm that specific hosts can e.g. host target plasmids, support conjugation, etc
Transformation	Transform DNA from select nodes into the selected (substrate) host cell.
incubation	Perform incubation procedure for select nodes. The ‘overnight incubation’ mode can simulate antibiotic screening
Extraction	Extract all plasmid DNA from selected nodes that are hosts (cells).
Gibson assembly designer	Design primers (based on user preferences) and apply Gibson assembly to DNA from selected nodes. Each source node must have only one DNA molecule. Where PCR products are used in a source node, the primer sequences will automatically be modified by extension to provide the correct overlap sequence
CRISPR digestion	Perform CRISPR digestion by using selected sgRNA and substrates