Background: Developed in 2008 by Collard and Mackill, SCoT markers target the flanking region around the start codon. Similar to RAPD, these use a single primer-based PCR amplification. SCoT primers are 18 nucleotides long and are designed to target the consensus sequence flanking start codon. The detection is based on a PCR assay and visualisation in ethidium bromide-stained agarose gel electrophoresis (Collard and Mackill, 2009b). These are dominant markers with locus specificity (Poczai et al., 2013).

Pros and cons: They have advantages like high polymorphism, simplified operation, inexpensive, reproducible, and simple primer design. However, reproducibility depends on factors such as primer length, annealing temperatures, and co-factors (Mg), which may influence reproducibility (Zeng et al., 2016).

Applications: SCoT has been applied in plant QTL mapping, bulked segregant analysis, and genetic diversity studies (Cabo et al., 2014; Gholami et al., 2021; Yeken et al., 2022). No study has been reported on farm animals in the public domain, but studies related to lower vertebrates such as sea cucumbers and fish are available. Abdelghany et al., 2023 describe the application of SCoT in sea cucumbers. SCoT was used to identify the genetic differentiation among five different species of sea cucumber in the Red Sea, Egypt, listed as Holothuria atra, H. impatiens, H. leucospilota, Actinopyga crassa, and A. mauritiana. Ten SCoT primers generated 150 amplicons with almost 52% polymorphism and 30 species-specific bands (Abdelghany et al., 2023).

Background: The limitations of RAPD and other random DNA markers demanded more specificity and reproducibility. In 2009, Collard and Mackill developed CDDP markers based on single primer-mediated amplification like RAPD. CDDP uses long primers that target plants’ genes responsive to abiotic and biotic stress (Collard and Mackill, 2009a; Golian et al., 2022; Sabo et al., 2022).

Pros and cons: CDDP markers overcome the reproducibility limitations of RAPD markers because primers are gene-specific, easy to apply and can be automated. However, occasional primer-related issues, such as primer length and high annealing temperatures, may arise that fail to ensure complete reproducibility (Poczai et al., 2013). CDDP markers are dominant, but no literature-based information exists for locus specificity.

Applications: CDDP was developed using sequences conserved within the plant genome responsive to abiotic and biotic stress. CDDP has its application in genetic analysis related to polymorphism, segregant analysis, and QTL mapping. No applications are available for these animal markers; however, many studies cited them for plant-related applications. Jiang and Zang have used CDDP to analyse genetic relationships in Rosa rugose. The study highlights the potential of this technology as a resource in germplasm conservation, analysing a plant’s genetic relatedness (Jiang and Zang, 2018).

Background: The technique for SRAP markers utilises two primers of 17-18 nucleotides in length. They target the open reading frame (ORF) of the gene. The primer structure determines the specificity of this technique, consisting of a core sequence of 13-14 bases, with the first 10-11 at the 5′ end. These bases have no specific constitution; these are termed filler sequences. They are followed by the sequence CCGG (targeting GC-rich ORF) in the forward primer and AATT (Targeting the AT-rich sequences) in the reverse primer. Three additional selective bases follow the core sequence at the 3′ end. It is important to note that the filler sequences for both primers must be different.

The protocol has a distinct PCR cycle. Initially, five cycles with an annealing temperature of 35°C, followed by 35 cycles of 50°C as the annealing temperature. It is resolved on a polyacrylamide gel. SRAP markers are dominant and locus-specific (Li and Quiros, 2001; Poczai et al., 2013).

Pros and cons: Performing SRAP assay is simple, reliable, has moderate throughput potential, and is cost-effective (Aneja et al., 2012). However, only 20% of the originally developed makers were co-dominant. Overlapping bands might lead to erroneous result interpretation. The 2-step PCR reaction makes it complex. The low annealing temperature might lead to non-specific primer annealing as well.

Application: Lin et al. (2003) constructed a genetic linkage map for cotton using the SRAP markers. The study used 136 different primer pairs and found 76 of these to be polymorphic. All the markers were evenly distributed among the linkage groups without loci clustering (Lin et al., 2003).

Alasaad et al. (2008) used SRAP to identify genetic variability among Fasciola using 10 SRAP primers. These Fasciola samples clustered into four groups but were unrelated to particular host species or the samples’ geographical locations. This study highlighted the application of SRAP for genetic diversity studies of parasites significant for human-animal health (Alasaad et al., 2008).

Background: In 2003, Hu and Vick developed TRAP using bioinformatics tools and EST database information to generate polymorphic markers around targeted gene sequences. Like SRAP, the method for TRAP relies on two primers of 18 nucleotides in length, except one is fixed and derived from EST, the other an arbitrary primer with an AT-rich (intron targeted) or GC-rich (exon targeted) core. It has two PCR cycles, one at 35°C for five cycles and another at 50°C. The bands are separated on a 6.5% polyacrylamide gel (Hu and Vick, 2003). TRAP is a dominant marker and is highly locus-specific.

Pros and cons: It has a simplified primer design step based on available sequence information. Samples do not require a pre-PCR treatment. However, slight mismatches are possible as the first five cycles are less stringent, and not all markers may linked to the gene of interest (Palumbo et al., 2007).

Applications: TRAP has been applied in mapping and tagging disease resistance traits in common beans., assessing the genetic diversity among sugarcane germplasm collections, and for marker-trait association studies (Alwala et al., 2006; Kumar et al., 2014; Miklas et al., 2006).

Background: CoRAP markers have been developed by combining principles of “SRAP” and “TRAP.” The method relies on a two-primer PCR amplification assay, with one arbitrary and one fixed primer. The fixed primer, similar to TRAP, is derived from the EST sequence of the target candidate genes, and the arbitrary primer is similar to the SRAP markers. It contains a conserved intron-specific sequence (CACGC). While the fixed primer provides specificity, the arbitrary primer promotes genome-wide accessibility (Wang et al., 2009). CoRAP are dominant markers and are locus-specific. The variation within the individual genomes leads to polymorphism, which can visualised on a polyacrylamide gel.

Pros and cons: CoRAP uses two primers, one derived from EST, which provides gene specificity. The other arbitrary primer ensures genome-wide accessibility and is more reliable than TRAP, easy to use, and reproducible) (Fabriki-Ourang and Yousefi-Azarkhanian, 2018; Wang et al., 2009).

Applications: CoRAP has been used to study the genetic relationship among 25 Salvia ecotypes/species using TRAP and CoRAP markers. Using a combination of 12 primers (four pairs of arbitrary primers and three pairs of fixed primers derived from EST sequences of Salvia miltorrhiza), 180 polymorphic loci (100%) data were obtained from this analysis, which helped the authors in clustering the 25 ecotypes/species into five major groups (Fabriki-Ourang and Yousefi-Azarkhanian, 2018).

Background: The PBA markers originate from the cytochrome P450 analogues, a monooxygenase enzyme conserved from animals to microbes, including plants. The enzyme plays a vital role in the biosynthesis of secondary metabolites and oxidative detoxification in plants, and it has a catabolic role in bacteria and oxidation of fatty acids and drug metabolism in animals (Behrendorff, 2021; Guengerich, 2021; Yamanaka et al., 2003). Yamanaka et al. (2003) developed these markers using PCR primers of mammalian P450 analogues in plants and used them to analyse intra and interspecific diversity within 51 plant species of 28 taxonomic families. The polymorphism can be observed on an agarose gel (Yamanaka et al., 2003).

Pros and cons: These functionally conserved genes are highly diverse in their genomic distribution. These are simple to use as universal primers exist. However, they may have PCR-related issues, resulting in technical failures (Poczai et al., 2013).

Application: Saini et al. (2013) used PBA markers to assess Moringa oleifera lam’s genetic variability. Seven pairs of PBA primers were used, and 40% polymorphism was identified. However, 48.68% and 48.57% polymorphism were reported for RAPD and ISSR primers, respectively. This study provides the practical applications of the PBA markers, where their role in genetic diversity and systematic breeding has been mentioned (Saini et al., 2013).

Background: TBP are markers based on the polymorphism within the introns of β-tubulins. Utilising the ubiquity of this gene, the primers designed for the first intron of different β-tubulin isotypes can reveal diverse fingerprints. The principle lies in the design of primers, targeting the difference in intron length. The polymorphism can be visualised on an agarose gel (Bardini et al., 2004). These are co-dominant markers but are not locus-specific (Morello et al., 2019).

Pros and cons: The method is simple and user-friendly, does not require prior sequence information for assay, and allows DNA barcoding. The intron is flanked by coding sequences on either side, enabling a more straightforward primer design. These are helpful when prior sequence information is unavailable (Bardini et al., 2004).

Applications: The markers have been applied to various plant species, such as Triticum, Camelina sativa, and grapes (Galasso et al., 2011; Gavazzi et al., 2016; Guadalupi et al., 2022). Gavazzi et al. (2016) used TBP as an alternative tool for genetically profiling grapes besides SSR. They could deduce comparable TBP results to six internationally recognised SSR markers (Gavazzi et al., 2016). Gianì et al. (2020) generated a genomic profile using TBP to authenticate animal species in meat and poultry. The study observed that each species has a distinct diagnostic fragment, which can assist in species-specific identification (Gianì et al., 2020).

Background: ITAP markers are based on polymorphism within the intron sequences (Poczai et al., 2013), such as using intron-exon splice junction targeted primers in identifying polymorphisms and their role in mapping polymorphic sites for various studies (Weining and Langridge, 1991). Xiong et al. (2013) developed ITAP based on the principles of arbitrary and EST-derived intron-exon splice junction primers. The method uses two primers, one targeting the intron-exon splice junction, the other derived as SRAP markers. ITAP are dominant markers, but no information is reported for loci specificity.

Pros and cons: Introns are sparsely distributed throughout the genome, and more events of genomic rearrangements occur within the introns than exons, a better indicator of polymorphism. The ITAP protocol is relatively simple and inexpensive and has been used for various applications.

Applications: ITAP markers have been used for various applications like genetic diversity assessment and sex identification of date palms and analysing the introgression of iron and zinc transfer-specific genes in Aegilops species (Atia et al., 2017a; Atia et al., 2017b; Xiong et al., 2013).

Background: A group of retro-transposons called short interspersed nuclear elements (SINEs) are short, repetitive, non-coding sequences, about 100–600 bp. SINEs are widely distributed throughout the genome and have various roles, such as genome organisation, evolution, and gene expression modulation (Kanhayuwa and Coutts, 2016). Therefore, these can be used as markers; the principle behind the method lies in the outward-facing primers that target the highly conserved SINE sequences. The variation within the amplicon product enables the polymorphism studies. These are dominant markers and locus-specific (Reiche et al., 2021; Seibt et al., 2012).

Pros and cons: ISAP markers are robust, cheaper than other markers such as AFLP, and can be easily automated through capillary electrophoresis. They do not require extensive laboratory equipment and can be analysed on an agarose gel. ISAP markers are more reproducible than RAPD, which provide similar informative banding. The directionality of change in ISAP can be determined, which cannot be done for SNP markers (Reiche et al., 2021; Seibt et al., 2012). ISAP marker development requires knowledge of the SINE sequences of the target species, and the variation among species is a significant limitation for wide-scale application.

Application: ISAP markers were developed to genotype potatoes; however, they have been translated for application in many other plant species. Sormin et al. used ISAP markers to genotype melon (Cucumis melo). The study includes the identification of melon-specific SINEs, and these melon-specific ISAP markers showed a higher polymorphism than potato-specific markers. The melon-specific ISAP markers were also tested in Coleus species to test their broader application. It was possible to differentiate between different Coleus accessions using melon-specific ISAPs (Sormin et al., 2021).

Background: SCAR is a genomic DNA fragment derived from a single locus identified by a pair of specific oligonucleotide primers. Paran and Michelmore first developed the markers in the 1990 s by cloning and sequencing locus-specific amplicons of RAPD markers. They designed 24-mer oligonucleotide primers using the sequence information from the single loci clone (Paran and Michelmore, 1993). SCAR is primarily a co-dominant locus-specific marker.

Pros and cons: SCAR overcomes the limitations of RAPD markers of reaction sensitivity and is more reliable than RAPD. The SCAR markers are specific to particular loci and can help develop specific markers for diagnostic or genotyping purposes (Amicucci et al., 1997; Paran and Michelmore, 1993). The only disadvantage of the technique is the laborious procedure. It is essential to clone and sequence the specific loci to design these primers.

Application: Yadav et al. (2012) developed SCAR markers to identify Bacopa monnieri from adulterants like Centella asiatica, Eclipta alba, and Malva rotundifolia. It could be cloned and sequenced using multiple RAPD primers and identifying plant-specific bands, and specific SCAR markers for Bacopa monnieri could be designed (Yadav et al., 2012). In another study, Didion et al. (2001) mapped a SCAR sequence to chromosome 6 from Porcine DNA, a fragment of 255 kb loci called DK122 (Didion et al., 2001).

Background: In Waugh et al. (1997) developed SSAPs to target transposable elements as markers. The SSAP markers are created by designing PCR primers that amplify the region between a priming site near the end of a transposable element and an adjacent restriction site flanking in the genomic DNA (Syed and Flavell, 2006). AFLP markers’ technique inspires the principle of SSAP but differs in the adaptor ligation, where a known sequence of Bare-1 specific radiolabeled primer was used as an adapter. These are dominant, locus-specific markers (Waugh et al., 1997).

Pros and cons: SSAPs are similar to the AFLP markers but identify retrotransposon insertion-mediated polymorphisms. These are highly informative, reliable, and efficient in detecting changes at individual and population levels (Jiang et al., 2016; Li et al., 2009). However, these markers require the genetic information of the retrotransposons of interest, which has limited its wide-scale application. Besides, it is a one-to 2-week process, making it laborious and cumbersome.

Applications: Berenyi et al. (2002) developed SSAP markers for the genetic analysis of sweet potatoes using the TY-1 copia retrotransposon. It has also been used in genotyping Vitis vinifera by creating SSAP markers for the Vine-1 retrotransposon (Berenyi et al., 2002; Labra et al., 2004).

Background: iSNAPs are small RNA-based molecular markers developed by Gui et al. (2011). They use the presence of small non-coding RNA of 20–24 nucleotides in length across the genome for marker development. The primer design explains these principles, which include mapping multiple small RNAs across the genome and designing primers at the two flanking ends of these multi-mapped small RNAs. The primers can be created within the small RNAs or their conserved 5′ and 3′ flanking regions (Gui et al., 2011).

Pros and cons: While the markers are reliable, offer high throughput, and are based on non-coding sequences (Poczai et al., 2013), they involve complex procedures and can be expensive regarding the sequencing costs associated with marker development.

Application: Other than the original study, no application studies were reported in the public domain. However, a few studies on related approaches which exploit miRNAs and other small RNA molecules in plant functional studies have been cited (Htwe et al., 2015; Solofoharivelo et al., 2014).

Background: PAAP are markers developed by Pang et al. (2009) using conserved promoter sequences of the cotton genome. They combined an RAPD primer and a degenerate promoter-specific primer to identify the polymorphism in the cotton genome and visualised it on a polyacrylamide gel (Pang et al., 2009).

Pros and cons: These are more reliable and informative than RAPD primers. However, they require the promoter information of the gene of interest, which may only be available for some species.

Applications: Mokate et al. (2017) used PAAP to assess divergence in Glycine max (Soybean). They observed that these markers produced significant polymorphic loci and highlighted the application of PAAP to study particular traits (Mokate et al., 2017). Table 2 lists the principles and PCR strategy for commonly used functional markers and their targets.

SNP is a single nucleotide substitution in genome sequences (Schork et al., 2000). Genome-wide distribution and automation amenability have made them the most preferred markers for humans, animals and plant genotyping studies (Zeng et al., 2017). SNPs have revolutionised marker-assisted studies in humans, animals and plants (Gao et al., 2023; Li et al., 2023; Pavlova et al., 2023; Raza et al., 2023). At the same time, there are many different ways of SNP genotyping, such as PCR, restriction digestion associated DNA sequencing, pyrosequencing and SNP arrays (Lawrie and Massey, 2023). SNP arrays are most popular, Various SNP arrays have been made for many organisms, including humans (Affymetrix Human SNP Array 6.0, Illumina HumanOmniZhongHua-8 Beadchip), dogs (Affymetrix Canine SNP Genotyping Array, Illumina CanineHD Beadchip), and cattle (Affymetrix Genome-Wide BOS 1 Bovine Array, Illumina BovineHD BeadChip) (Liu et al., 2017). A large number of GWAS studies have been conducted using these SNP chips. SNPs are widely used in various domains of biological sciences, which could be difficult to accommodate here. Therefore, a brief description of the different SNP arrays and their application in humans, animals, and plants is provided in the Supplementary Material.

Repetitive DNA sequences are the homologous DNA sequences found in multiple genome copies. Their genome-wide occurrence makes them a potential marker. They can be grouped based on occurrences such as highly repetitive (Satellite DNA) and moderately repetitive DNA sequences (Tandem and interspersed repeat sequences) (Pelley, 2007). DNA tandem repeats can be further categorised into microsatellites, minisatellites, and megasatellites (Trent, 2012), whereas interspersed repeat sequences are further classified into RNA and DNA transposons-based markers. Repetitive DNA markers offer the advantages of co-dominance and genome-wide polymorphism coverage. It is widely used in plant and animal genotyping studies (Bora et al., 2023; Lahkar et al., 2023; Liu et al., 2023; Loukovitis et al., 2023; Mauger et al., 2023; Yeo et al., 2023). The scope of the current review is limited to the random and functional markers. Therefore, only a brief description of the marker and the application of each repetitive DNA marker is provided in the Supplementary Material.