Elsevier

Genomics

Volume 24, Issue 2, 15 November 1994, Pages 225-233
Genomics

Regular Article
Automation of Genetic Linkage Analysis Using Fluorescent Microsatellite Markers

https://doi.org/10.1006/geno.1994.1610Get rights and content

Abstract

Automation of the typing of genetic markers offers advantages of speed, accuracy, and cost in the mapping of genetic traits and the construction of high-resolution linkage maps. We have developed an automated linkage analysis system by (i) using a robotic pipettor to set up polymerase chain reactions (PCR) to amplify microsatellites with incorporation of a single fluorescent label; (ii) using an automated sequencing apparatus for detection of the PCR products; (iii) sizing alleles automatically by the use of internal and external standards; (iv) iteratively filtering out nonallelic fragments and checking for Mendelian consistency; (v) calculating the probabilities of selected genotypes; and (vi) automatically formatting the results for input to linkage analysis programs. The method provides accurate sizing of alleles, minimizes the risk of error during manual reading and transcription of data, and increases the throughput of reliable data. It brings any inconsistencies or ambiguities in the data to the attention of the user and facilitates examination of the raw data. The ALF/ALP system, together with new, optimized microsatellite sets, particularly tetranucleotide repeats, is likely to be well-suited to fully automatic genetic linkage analysis.

References (0)

Cited by (68)

  • Microsatellite and mitochondrial DNA markers unveil the genetic structure of Nellore Palla sheep of India

    2022, Small Ruminant Research
    Citation Excerpt :

    Capillary electrophoresis is advantageous over agarose gel electrophoresis due to its ability to measure the size of PCR products with very high resolution (WHO (World Health Organization) and the Medicines for Malaria Venture, 2007; Liljander et al., 2009). Fluorescence-based microsatellite loci detection and allele scoring on an automated DNA fragment analyser is one of the most accurate methods for genotyping as overlapping alleles are co-separated by labelling with different fluorophores and multi-pooling (Mansfield et al., 1994; Heller, 2001). The value observed over the range of loci in the present study could be regarded as a reasonable indicator of genetic variation within the populations (MacHugh et al., 1997).

  • A multiplex microsatellite PCR method for evaluating genetic diversity in grass carp (Ctenopharyngodon idellus)

    2018, Aquaculture and Fisheries
    Citation Excerpt :

    The use of capillary electrophoresis with fluorescently labelled primers provides high detection sensitivity of amplified DNA fragments (Huang, Huang, Hu, & Chang, 2006; Ramachandran et al., 2003; Shi et al., 2003). Multiplex SSR-PCR as an effective, economical and accurate approach for genotyping and germplasm evaluation, it enhances the visualization of polymorphic DNA fragments and was the excellent option of constructed the method for evaluating genetic information of grass carp (Archak, Lakshminarayanareddy, & Nagaraju, 2007; Mansfield et al., 1994; Ziegle et al., 1992). In this study, we constructed a multiplex SSR-PCR detection method possessed high polymorphism and heterozygosity.

  • Guidelines for the selection of functional assays to evaluate the hallmarks of cancer

    2016, Biochimica et Biophysica Acta - Reviews on Cancer
    Citation Excerpt :

    SNP arrays are capable of detecting copy-neutral loss of heterozygosity (so-called uniparental disomy or gene conversion) and distinguish alleles at specific polymorphic sites [189]. To detect microsatellite instability (MSI), the gold standard approach amplifies known microsatellite regions by PCR and quantifies the lengths of PCR products by gel or capillary electrophoresis using autoradiography, silver staining [190] or fluorescent methods [191,192]. The microsatellite length of the sample is compared to normal DNA to determine MSI status.

  • Multiplex SSR-PCR approaches for semi-automated genotyping and characterization of loci linked to blast disease resistance genes in rice

    2015, Comptes Rendus - Biologies
    Citation Excerpt :

    The use of capillary electrophoresis with fluorescently labelled primers provides high detection sensitivity of amplified DNA fragments [33–35]. This procedure is an effective, economical and accurate approach for genotyping, germplasm evaluation and enhances the visualisation of polymorphic DNA fragments, representing multiple loci from across the genome [36–38]. Nowadays, this method has been developed and used for several plant species, such as Oryza sativa [39–41].

View all citing articles on Scopus
View full text