In a Q & A session of the AGH635 lecture, one of the student asked the following question: “Was the term loci and alleles also applicable when one discussed about ISSR (Inter Simple Sequence Repeat) marker? What were the loci and what were the allele in the ISSR marker analysis?”
Let me just remind you the previous posting, the term locus (pl.: loci) was a common terminology associated with a location in the genome of an organism. In the Mendelian genetics, the term of a locus controlling flower color clearly described a location somewhere in the genome associated with gene(s) controlling the flower color character. Meanwhile, the term allele was associated with alternative gene products resided in certain locus. In the above example, a gene product controlling a red flower color was an allele of flower color locus. Alternatively, the other allele for this locus was the gene product controlling a white color one.
Similar to RAPD marker, ISSR markers were simply a PCR amplified DNA fragments generated using a simple sequence repeat (SSR) associated sequence as oligo-nucleotide primer and genomic DNA as template (see Figure 1).
In a single PCR reaction using certain SSR associated sequence primer, there could be a number of different sizes of amplified DNA fragments. A certain size of the PCR amplified fragment using a single SSR associated sequence primer was generated from certain location in the genome. Hence, different sizes of the PCR amplified fragments were generated from different locations in the genome (Figure 2).
When one analyzes ISSR marker for a number of individuals in a population using a single SSR associated sequence primer and one looks at certain size of PCR amplified product (i.e. Fragment #2), there will be two possibilities. For one individual, this amplified product might be present while for the other it might be absent.
Back to the questions – one of the AGH635 student’s asked, what one regards as locus in an ISSR marker analysis was usually named after the primer used and the amplified product sizes, just like in RAPD marker. In the Figure 2, PCR amplification of the plants using a single SSR associated sequence primer resulted in three different sizes of amplified products. Hence, three loci were identified in this ISSR analysis.
In ISSR analysis – one PCR reaction using a single SSR associated sequence primer could generate a number of markers. Therefore, the ISSR marker was categorized as multi-loci markers
Just like in RAPD marker, the rule in naming of the locus in ISSR markers analysis were: “Name of the primer” used to amplify the marker and followed by “the sizes of the PCR amplified products.” If one looks at a particular locus and survey the amplified product across samples, then the PCR amplified product is either present or absent. In ISSR marker analysis, the present and absence of amplified product for a locus analyzed represent the alternative alleles for that locus. Therefore, there were only two possible alleles for each locus in the ISSR analysis, such as: present and absence of PCR amplified product.
To summarize, in ISSR marker analysis – the locus is defined as the name of the primer, followed by the size of amplified product. There are only two alternative alleles for each locus in ISSR marker, such as PCR product is either present or absence.