Limitations on the use of polymerase chain reaction – restriction fragment length polymorphism analysis of the rDNA NTS2 region for the taxonomic classification of the speciesSaccharomyces cerevisiae

2005 ◽  
Vol 51 (9) ◽  
pp. 759-764 ◽  
Author(s):  
Andrea Pulvirenti ◽  
Lisa Solieri ◽  
Luciana De Vero ◽  
Paolo Giudici
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Polymerase Chain Reaction ◽  
Fragment Length Polymorphism ◽  
Rflp Analysis ◽  
Chain Reaction ◽  
D2 Domain ◽  
Pcr Rflp ◽  
Its Regions ◽  
Polymerase Chain ◽  
Restriction Fragment Length

Different molecular techniques were tested to determine which was the most effective in the identification of Saccharomyces cerevisiae strains. In particular, polymerase chain reaction – restriction fragment length polymorphism (PCR–RFLP) analysis of the internal transcribed spacer (ITS) regions and the nontranscribed spacer 2 (NTS2) region, sequencing of the D1/D2 domain, and electrophoretic karyotyping were applied to 123 yeast strains isolated from different sourdoughs and tentatively attributed to the species S. cerevisiae. All of the strains tested showed an identical PCR–RFLP pattern for the ITS regions, an identical nucleotide sequence of the D1/D2 domain, and the typical electrophoretic karyo type of S. cerevisiae. In contrast, 14 out of the 123 strains tested showed some polymorphism with BanI restriction analysis of the NTS2 region. Our results indicate that while the sequencing of the D1/D2 domain, the PCR–RFLP analysis of the ITS regions, and the electrophoretic karyotype can be employed successfully to identify S. cere visiae strains, PCR–RFLP analysis of the NTS2 region does not allow a consistent and accurate grouping for S. cere visiae strains. The fact that the NTS2 region of a small number of strains (8.78% of the total strains tested) is different from that of the other S. cerevisiae strains confirms that molecular methods should always be tested on a great number of strains.Key words: ribosomal DNA, Saccharomyces cerevisiae, yeast identification.

scholarly journals Identifikasi Keragaman Gen DGAT1 serta Asosiasinya terhadap Karakteristik Karkas dan Sifat Perlemakan Domba

2019 ◽  
Vol 6 (2) ◽  
pp. 259
Author(s):  
Asep Gunawan ◽  
Ratna Sholatia Harahap ◽  
Kasita Listyarini ◽  
Cece Sumantri
Keyword(s):  
Polymerase Chain Reaction ◽  
Single Nucleotide Polymorphism ◽  
Fragment Length Polymorphism ◽  
Nucleotide Polymorphism ◽  
Chain Reaction ◽  
Single Nucleotide ◽  
Dgat1 Gene ◽  
Pcr Rflp ◽  
Polymerase Chain ◽  
Restriction Fragment Length

ABSTRAK Karakteristik karkas dan sifat perlemakan pada daging domba dikontrol oleh banyak gen salah satunya gen DGAT1 (Diacylglycerol Acyltransferasel 1). Penelitian ini bertujuan mengidentifikasi SNP (Single Nucleotide Polymorphism) gen DGAT1 pada titik mutasi g.8539 C>T dan asosiasinya terhadap karakteristik karkas dan sifat perlemakan pada domba Indonesia. Total sampel domba yang digunakan sebanyak 150 buah terdiri dari 35 sampel domba compass agrinak (DCA), 36 sampel domba barbados cross (DBC), 41 sampel domba komposit garut (DKG), 20 sampel domba ekor gemuk (DEG), dan 18 sampel domba ekor tipis (DET). Karakteristik karkas dan sifat perlemakan diukur dari domba jantan berumur 10-12 bulan. Identifikasi keragaman DGAT1|ALuI dianalisis dengan metode PCR-RFLP (Polymerase Chain Reaction-Restriction Fragment Length Polymorphism). Hasil keragaman gen DGAT1 bersifat polimorfik dalam DET dan DEG, sedangkan DCA, DBC, dan DKG bersifat monomorfik. Dua genotipe disebut CC dan  CT ditemukan dalam DET dan DEG. Titik mutasi gen DGAT1 berasosiasi (P<0.05) dengan karakteristik karkas, yaitu bobot dan panjang karkas. Selain itu, keragaman gen DGAT1 juga berasosiasi signifikan (P<0.05) dengan asam lemak jenuh, yaitu asam stearat (C18:0) dan asam arakidat (C20:0) dan asam lemak tak jenuh tunggal, yaitu asam oleat (C18:1n9c). Gen DGAT1 memiliki kontribusi dalam karakteristik karkas dan komposisi asam lemak pada domba.Kata Kunci: domba, gen DGAT1, karakteristik karkas, PCR-RFLP, sifat perlemakan                                                              ABSTRACT            Characteristic of carcass and fatness traits of sheep is regulated by many genes such as DGAT1 (Diacylglycerol Acyltransferasel 1) gene. The research was aimed to investigate SNP (Single Nucleotide Polymorphism) of DGAT1 and its association with characteristic of carcass and fatness traits in Indonesian sheep. A total sample of sheeps used 150 rams of 10–12 months consisted 35 samples of compas agrinak sheep (CAS), 36 of barbados cross (BCS), 41 of garut composite (GCS), 20  of javanese fat tailed (JFT), and 18 of javanese thin tailed (JTT). Identification variant of DGAT1|ALuI were performed by PCR-RFLP (Polymerase Chain Reaction-Restriction Fragment Length Polymorphism). The results of polymorphism of DGAT1 were found in JTT and JFT. However, SNP of DGAT1 in CAS, BCS and GCS were monomorfic. Two genotype namely CC and CT were found in JTT and JFT populations. A SNP of the DGAT1 was associated (P<0.05) with characteristic of carcass, including weight and length of carcass. The variant of DGAT1 was associated too with saturated fatty acids (SFA) including stearic acid (C18:0) and arachidic acid (C20:0), and mono unsaturated fatty acid (MUFA) including oleic acid (C18:1n9c). The DGAT1 gene was contribute to characteristic carcass and fatty acid composition in sheep.Keywords: DGAT1 gene, characteristic carcass, fatness traits, PCR-RFLP, sheep

Genetic analysis of nuclear DNA restriction fragment patterns

Genome ◽  
1991 ◽  
Vol 34 (5) ◽  
pp. 693-703 ◽  
Author(s):  
Elizabeth M. Gillet
Keyword(s):  
Polymerase Chain Reaction ◽  
Restriction Fragment Length Polymorphism ◽  
Genetic Analysis ◽  
Restriction Fragment ◽  
Length Polymorphism ◽  
Fragment Length Polymorphism ◽  
Rflp Analysis ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Restriction Fragment Length

Restriction fragment length polymorphism (RFLP) analysis in the broad sense is the analysis of differences in restriction fragment pattern produced by defined target segments within or between cell compartments, cell types, etc., in a single individual or in different individuals. Thus both molecular hybridization and DNA amplification by two-primer extension using the polymerase chain reaction can define target segments for RFLP analysis. The two techniques are outlined with special consideration of characteristics important for genetic analysis. The mode of inheritance of restriction fragment patterns as a prerequisite for their use as genetic markers in inheritance studies is explained, leading to criticism of common usage. The importance of internal restriction sites for the determination of allelic variation is stressed. It is shown that, if target segments are under the control of a single nuclear diploid restriction fragment locus, then complete reconstruction of all parental target segments requires controlled crosses between individuals of like restriction fragment pattern.Key words: genetic analysis, inheritance, restriction fragment length polymorphism, controlled cross, polymerase chain reaction.

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