AFLP fingerprinting for paternity testing in ducks
TL;DR: AFLP fingerprinting might be a suitable method for duck paternity testing and shows high reproducibility, from the family testing, which found that the fingerprints of all the offspring were derived from one or other parent.
Abstract: 1. The accuracy and reproducibility of AFLP fingerprinting was investigated in the duck (Anas Platyrhynchos), using a multicolour fluorescent labeling technique. The fluorescent labelling fragments were separated on a capillary electrophoresis-base ABI PRISM 3100 Genetic Analyzer. 2. A total of 337 AFLP peaks with 103 of them being polymorphic markers were generated by 16 sets consisting of EcoRI/TaqI primer pair combinations. The number and size range of AFLP polymorphisms detected per primer pair varied from 3 to 11 and 58 to 290 bp, respectively. About 30.6% (103/337) of AFLP peaks were detected polymorphisms, with an average of 6.4 polymorphic markers per primer pair. 3. The clear polymorphic peaks were amplified with EcoR+AC/Taq+AC primer combinations. The AFLP peaks showed high reproducibility. From the family testing, we found that the fingerprints of all the offspring were derived from one or other parent. Therefore, we conclude that AFLP fingerprinting might be a suitable method for duck paternity testing.
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TL;DR: This study is the first to introduce a PCR-melting curve analysis (PCR/MCA) to identify the gender of birds by genomic DNA, which is gel-free, quick, and inexpensive and holds a great potential for use in high throughput sexing of other avian species, as well.
Abstract: Combination of CHD (chromo-helicase-DNA binding protein)-specific polymerase chain reaction (PCR) with electrophoresis (PCR/electrophoresis) is the most common avian molecular sexing technique but it is lab-intensive and gel-required. Gender determination often fails when the difference in length between the PCR products of CHD-Z and CHD-W genes is too short to be resolved. Here, we are the first to introduce a PCR-melting curve analysis (PCR/MCA) to identify the gender of birds by genomic DNA, which is gel-free, quick, and inexpensive. Spilornis cheela hoya (S. c. hoya) and Pycnonotus sinensis (P. sinensis) were used to illustrate this novel molecular sexing technique. The difference in the length of CHD genes in S. c. hoya and P. sinensis is 13-, and 52-bp, respectively. Using Griffiths' P2/P8 primers, molecular sexing failed both in PCR/electrophoresis of S. c. hoya and in PCR/MCA of S. c. hoya and P. sinensis. In contrast, we redesigned sex-specific primers to yield 185- and 112-bp PCR products for the CHD-Z and CHD-W genes of S. c. hoya, respectively, using PCR/MCA. Using this specific primer set, at least 13 samples of S. c. hoya were examined simultaneously and the Tm peaks of CHD-Z and CHD-W PCR products were distinguished. In this study, we introduced a high-throughput avian molecular sexing technique and successfully applied it to two species. This new method holds a great potential for use in high throughput sexing of other avian species, as well.
62 citations
Cites methods from "AFLP fingerprinting for paternity t..."
...This limitation has been overcome by some methods such as redesigned PCR primers [18] and our proposed approach, as well as PCR-RFLP [23,25], RAPD [26] and AFLP [27] fingerprintings....
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TL;DR: From this study, it is suggested to use a pair of 2550F and 2718R primers for distinguishing a male from a female bird.
Abstract: Visually identifying the sex of a bird can be difficult. It cannot be done in half the world's species when they are adults, and virtually none can be sexed as chicks. Despite this, the sex of a bird is vital for captive breeding. An increased number of birds are being sexed using DNA amplification techniques. In this approach, the CHD-W and CHD-Z are distinguished by the amplification of an intron present in both genes. PCR products on the gel electrophoresis vary in size revealing one band in males at the CHD-Z, and two bands in females corresponding to both the CHD-W and CHD-Z. Two independent sets of primer (P8/P2 and 2550F/2718R) were used to amplify the CHD gene region from both the Z and W chromosome. One hundred and ten (110) birds were sexed using first pair of primers: (P8/P2). Sexing results indicated that 81.8% were successfully determined, 12.7% failed to be amplified and 5.5% were not perfectly determined because the PCR products showed thick band. The thick band caused misidentified female to male birds. An alternative primer (2550F/2718R) was applied to solve the problem. Two hundreds and twenty-nine birds were sexed and the results showed 100% successfully determined. From this study, it is suggested to use a pair of 2550F and 2718R primers for distinguishing a male from a female bird.
26 citations
TL;DR: MCA using proposed primer sets was a robust gender identification method for the three Columbidae species tested and females contained two Tm peaks, whereas males contained one.
19 citations
TL;DR: This AFLP linkage map provides important information for establishing a duck chromosome map, for mapping quantitative trait loci (QTL mapping) and for breeding applications.
Abstract: Amplified fragment length polymorphism (AFLP) with multicolored fluorescent molecular markers was used to analyze duck (Anas platyrhynchos) genomic DNA and to construct the first AFLP genetic linkage map. These markers were developed and genotyped in 766 F2 individuals from six families from a cross between two different selected duck lines, brown Tsaiya and Pekin. Two hundred and ninety-six polymorphic bands (64% of all bands) were detected using 18 pairs of fluorescent Taq I/Eco RI primer combinations. Each primer set produced a range of 7 to 29 fragments in the reactions, and generated on average 16.4 polymorphic bands. The AFLP linkage map included 260 co-dominant markers distributed in 32 linkage groups. Twenty-one co-dominant markers were not linked with any other marker. Each linkage group contained three to 63 molecular markers and their size ranged between 19.0 cM and 171.9 cM. This AFLP linkage map provides important information for establishing a duck chromosome map, for mapping quantitative trait loci (QTL mapping) and for breeding applications.
16 citations
Cites background from "AFLP fingerprinting for paternity t..."
...[8], several studies have shown that AFLP markers follow Mendelian inheritance rules and that the technique is highly reproducible, powerful and efficient [9]....
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TL;DR: The MCA-based real-time PCR combined with the proposed primer redesign provided a high-throughput method of identifying sex in C. chinensis and G. gallus, which was useful for biomedical research and hatcheries.
16 citations
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TL;DR: The AFLP technique provides a novel and very powerful DNA fingerprinting technique for DNAs of any origin or complexity that allows the specific co-amplification of high numbers of restriction fragments.
Abstract: A novel DNA fingerprinting technique called AFLP is described. The AFLP technique is based on the selective PCR amplification of restriction fragments from a total digest of genomic DNA. The technique involves three steps: (i) restriction of the DNA and ligation of oligonucleotide adapters, (ii) selective amplification of sets of restriction fragments, and (iii) gel analysis of the amplified fragments. PCR amplification of restriction fragments is achieved by using the adapter and restriction site sequence as target sites for primer annealing. The selective amplification is achieved by the use of primers that extend into the restriction fragments, amplifying only those fragments in which the primer extensions match the nucleotides flanking the restriction sites. Using this method, sets of restriction fragments may be visualized by PCR without knowledge of nucleotide sequence. The method allows the specific co-amplification of high numbers of restriction fragments. The number of fragments that can be analyzed simultaneously, however, is dependent on the resolution of the detection system. Typically 50-100 restriction fragments are amplified and detected on denaturing polyacrylamide gels. The AFLP technique provides a novel and very powerful DNA fingerprinting technique for DNAs of any origin or complexity.
12,960 citations
"AFLP fingerprinting for paternity t..." refers background or methods in this paper
...The main consideration for methods based on DNA fingerprinting of a defined molecular marker is a high degree of reproducibility (Vos et al., 1995)....
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...The DNA fingerprinting by the AFLP method was originally developed by Vos et al. (1995)....
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...Vos et al. (1995) reported that selective PCR primers which had three nucleotides added on to their 30 ends generated a manageable number of polymorphic peaks from mammalian genomic DNA....
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...The AFLP fingerprinting technique depends on PCR amplification of a subset of DNA restriction-ligation fragments (Vos et al., 1995)....
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...Previous experiments suggested that a wide range concentration of genomic DNA could be successfully employed in the AFLP technique (Vos et al., 1995)....
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TL;DR: A new marker type, named SNP, for Single Nucleotide Polymorphism, is now on the scene and has gained high popularity, even though it is only a bi-allelic type of marker.
Abstract: During the last ten years, the use of molecular markers, revealing polymorphism at the DNA level, has been playing an increasing part in animal genetics studies. Amongst others, the microsatellite DNA marker has been the most widely used, due to its easy use by simple PCR, followed by a denaturing gel electrophoresis for allele size determination, and to the high degree of information provided by its large number of alleles per locus. Despite this, a new marker type, named SNP, for Single Nucleotide Polymorphism, is now on the scene and has gained high popularity, even though it is only a bi-allelic type of marker. In this review, we will discuss the reasons for this apparent step backwards, and the pertinence of the use of SNPs in animal genetics, in comparison with other marker types.
1,010 citations
"AFLP fingerprinting for paternity t..." refers methods in this paper
...A review on SNP and other types of molecular markers and their use in animal genetics....
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...The AFLPs are bi-allelic dominant markers and according to the method of analysis they can be scored co-dominantly (Ajmone-Marsan et al., 1997; Herbergs et al., 1999; Vignal et al., 2002)....
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...There are many different types of sequence variation and they are revealed by molecular biology techniques which include amplified fragment length polymorphisms (AFLP), random amplified polymorphic DNA (RAPD), restriction fragment length polymorphism (RFLP), microsatellite, minisatellite, simple sequence repeats (SSRs) and single nucleotide polymorphism (SNP) (Vignal et al., 2002)....
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...…biology techniques which include amplified fragment length polymorphisms (AFLP), random amplified polymorphic DNA (RAPD), restriction fragment length polymorphism (RFLP), microsatellite, minisatellite, simple sequence repeats (SSRs) and single nucleotide polymorphism (SNP) (Vignal et al., 2002)....
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TL;DR: Because of their high replicability and ease of use, AFLP markers have emerged as a major new type of genetic marker with broad application in systematics, pathotyping, population genetics, DNA fingerprinting and quantitative trait loci (QTL) mapping.
Abstract: Amplified fragment length polymorphisms (AFLPs) are polymerase chain reaction (PCR)-based markers for the rapid screening of genetic diversity. AFLP methods rapidly generate hundreds of highly replicable markers from DNA of any organism; thus, they allow high-resolution genotyping of fingerprinting quality. The time and cost efficiency, replicability and resolution of AFLPs are superior or equal to those of other markers [allozymes, random amplified polymorphic DNA (RAPD), restriction fragment length polymorphism (RFLP), microsatellites], except that AFLP methods primarily generate dominant rather than co-dominant markers. Because of their high replicability and ease of use, AFLP markers have emerged as a major new type of genetic marker with broad application in systematics, pathotyping, population genetics, DNA fingerprinting and quantitative trait loci (QTL) mapping.
992 citations
"AFLP fingerprinting for paternity t..." refers background in this paper
...Numerous (>1000) independent genetic loci can be rapidly screened by the AFLP technique, which underlie the genetic diversity and variation distributed randomly in genomic sequences of organisms (Mueller and Wolfenbarger, 1999)....
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TL;DR: Comparison of the newly obtained data with results previously obtained by well-established genotypic and chemotaxonomic methods shows the superior discriminative power of AFLP towards the differentiation of highly related bacterial strains that belong to the same species or even biovar (i.e. to characterize strains at the infrasubspecific level).
Abstract: We investigated the usefulness of a novel DNA fingerprinting technique, AFLP, which is based on the selective amplification of genomic restriction fragments by PCR, to differentiate bacterial strains at the subgeneric level. In total, 147 bacterial strains were subjected to AFLP fingerprinting: 36 Xanthomonas strains, including 23 pathovars of Xanthomonas axonopodis and six pathovars of Xanthomonas vasicola, one strain of Stenotrophomonas, 90 genotypically characterized strains comprising all 14 hybridization groups currently described in the genus Aeromonas, and four strains of each of the genera Clostridium, Bacillus, Acinetobacter, Pseudomonas and Vibrio. Depending on the genus, total genomic DNA of each bacterium was digested with a particular combination of two restriction endonucleases and the resulting fragments were ligated to restriction halfsite-specific adaptors. These adaptors served as primer-binding sites allowing the fragments to be amplified by selective PCR primers that extend beyond the adaptor and restriction site sequences. Following electrophoretic separation on 5% (w/v) polyacrylamide/8.3 M urea, amplified products could be visualized by autoradiography because one of the selective primers was radioactively labelled. The resulting banding patterns, containing approximately 30-50 visualized PCR products in the size range 80-550 bp, were captured by a high-resolution densitoscanner and further processed for computer-assisted analysis to determine band-based similarity coefficients. This study reveals extensive evidence for the applicability of AFLP in bacterial taxonomy through comparison of the newly obtained data with results previously obtained by well-established genotypic and chemotaxonomic methods such as DNA-DNA hybridization and cellular fatty acid analysis. In addition, this study clearly demonstrates the superior discriminative power of AFLP towards the differentiation of highly related bacterial strains that belong to the same species or even biovar (i.e. to characterize strains at the infrasubspecific level), highlighting the potential of this novel fingerprinting method in epidemiological and evolutionary studies.
561 citations
"AFLP fingerprinting for paternity t..." refers background in this paper
...The restriction enzymes EcoRI/MseI, HindIII/MseI and Apa1/TaqI have been shown to be the most suitable enzymes to digest genomes which have poor, 40—50%, or rich GþC nucleotide content respectively (Janssen et al., 1996)....
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TL;DR: A review of research areas in the study of wild species of animals where the AFLP method should be a very valuable tool to help molecular ecologists to identify when AFLP is likely to be superior to other more established methods, such as microsatellites, SNP (single nucleotide polymorphism) analyses and multigene DNA sequencing.
Abstract: Researchers in the field of molecular ecology and evolution require versatile and low-cost genetic typing methods. The AFLP (amplified fragment length polymorphism) method was introduced 10 years ago and shows many features that fulfil these requirements. With good quality genomic DNA at hand, it is relatively easy to generate anonymous multilocus DNA profiles in most species and the start-up time before data can be generated is often less than a week. Built-in dynamic, yet simple modifications make it possible to find a protocol suitable to the genome size of the species and to screen thousands of loci in hundreds of individuals for a relatively low cost. Until now, the method has primarily been applied in studies of plants, bacteria and fungi, with a strong bias towards economically important cultivated species and their pests. In this review we identify a number of research areas in the study of wild species of animals where the AFLP method, presently very much underused, should be a very valuable tool. These aspects include classical problems such as studies of population genetic structure and phylogenetic reconstructions, and also new challenges such as finding markers for genes governing adaptations in wild populations and modifications of the protocol that makes it possible to measure expression variation of multiple genes (cDNA-AFLP) and the distribution of DNA methylation. We hope this review will help molecular ecologists to identify when AFLP is likely to be superior to other more established methods, such as microsatellites, SNP (single nucleotide polymorphism) analyses and multigene DNA sequencing.
491 citations
"AFLP fingerprinting for paternity t..." refers background in this paper
...…study sex determination (Horng and Huang, 2003; Huang et al., 2003; Horng et al., 2006), genetic relationships (Minvielle et al., 2000), classification (Cervera et al., 2005), molecular evolution (Laval et al., 2002), population ecology (Bensch and Akesson, 2005) and phylogeny (Laval et al., 2000)....
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...AFLP analysis requires comparatively short start-up time in most species and has the potential to detect the polymorphisms of PCR products (Bensch and Akesson, 2005)....
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