AutoCloner

AutoCloner is a custom pipeline that designs homeologue-specific primers for polyploids. It achieves this by extracting SNP locations unique to your gene of interest, and placing the 3` ends of primers at these SNPs.

Whilst there is an existing tool for designing KASP primers in wheat, PolyMarker (Ramirez-Gonzalez et al., 2015), there is not currently anything available for researchers who wish to clone entire genes. This is where AutoCloner comes in.

Please input your sequence in the form to the right. AutoCloner will design a stacked PCR arrangement of primers depending on your specified minimum and maximum product lengths.

Source code and documentation at GitHub.

Fig. 1 Illustration of the AutoCloner pipeline. Flanking regions are extracted from the genome before mining for SNPs and subsequent primer design.

Sequence name:
Genome:
Additional genomes:Integrate genomes of other wheat varieties (Clavijo et al., 2017) into the pipeline. This can increase the fidelity of SNPs at the cost of a longer computation time. This setting will be ignored for genomes other than bread wheat.	Paragon: Claire: Robigus: Cadenza:
Sequence: AutoCloner works best with genomic sequences, but will also work with a CDS

	Min product length:Specify the minimum length of the desired PCR product Max product length:Specify the maximum length of the desired PCR product
	Start buffer:The number of flanking nucleotides to extract upstream of the input sequence End buffer:The number of flanking nucleotides to extract downstream of the input sequence
	Single product:Return a primers for a single PCR product rather than a stacked arrangement? Mask inter-HSP distances:Mask regions between BLAST hits? These could affect the multiple alignment software negatively
Alignment method: Muscle is a good general purpose alignment software; Dialign uses anchors to inform the alignment - this is useful when large numbers of homologous regions of different lengths are present
CDS max intron size: The threshold distance under which to group BLAST hits together

Use own multiple sequence alignment

Based at the University of Bristol with support from BBSRC.

AutoCloner maintained by Alex Coulton.