DNA Analysis
Steven H. Y. Wong, Iraving Sunshine in Handbook of Analytical Therapeutic Drug Monitoring and Toxicology, 2017
A second commercial dot blot kit, known as the AmpliType® PM PCR Amplification and Typing Kit, involves as additional five genetic loci and collectively raises the discriminatory power to one in a few thousand.51 The five different loci simultaneously tested are: glycophorin A (GYPA), hemoglobin gamma-globin chain (HBGG), group specific component (GC), D7S8, and low density lipoprotein receptor (LDLR). Glycophorin A is the protein that carries the classical MN blood-group polymorphism. The GC (group specific component) is a human serum protein polymorphism that has been typed for years using protein chemistry methods. The low density lipoprotein receptor (LDLR), D7S8, and HB γ-globin (HBGG) loci have not previously been utilized in forensic work. The single amino acid difference polymorphism in the γ-chains of human hemoglobin F, resulting from apparently duplicated γ-chain genes, was described long ago. AmpliType® PM detects the basis for this difference at the DNA level. Each of these systems exhibits two or three alleles, resulting in three to six genotypes.
ChIP-seq analysis
Altuna Akalin in Computational Genomics with R, 2020
There are four steps in ChIP quality control: Sample correlation clustering: Clustering of the pair-wise correlations between genome-wide signal profiles.Data visualization in a genomic browser.Average fragment length determination: Determining whether the ChIP was enriched for fragments of a certain length.Visualization of GC bias. Here we will plot the ChIP enrichment versus the average GC content in the corresponding genomic bin.
Cryptosporidium
Dongyou Liu in Laboratory Models for Foodborne Infections, 2017
The genome sequences of C. parvum IOWA, C. hominis TU502, and C. muris were released on CryptoDB in 2003 and 2011, respectively [11–13]. Composed of eight chromosomes each, these genomes are of 9.1–92 Mb in size, with 26%–30% GC content and 94%–97% nucleotide identity. Altogether, about 4000 protein-encoding genes are present. Cryptosporidium genomes appear to be unusual among eukaryotes in having a degenerate “mitosome” (located in the posterior end of sporozoites) instead of a mitochondrion, with accompanied loss of many mitochondrial proteins, including those required for the TCA cycle, oxidative phosphorylation, and fatty acid oxidation as well as de novo biosynthesis of amino acids, nucleotides, and sugars. These gene losses render Cryptosporidium species heavily reliant on scavenging nutrients from the host rather than de novo biosynthesis [12–15]. In addition, sequencing analyses of C. hominis IbA10G2 (the most virulent subtype responsible for all outbreaks in Europe and Australia) and IaA28R4 (a dominant outbreak subtype in the United States) uncovered major differences in the 5′ and 3′ ends of chromosome 6 and the putative virulence determinant gp60 region, suggesting that genetic recombination plays a potential role in the emergence of hypertransmissible C. hominis subtypes [16,17].
Comparative genome analysis of Alkhumra hemorrhagic fever virus with Kyasanur forest disease and tick-borne encephalitis viruses by the in silico approach
Published in Pathogens and Global Health, 2018
Navaneethan Palanisamy, Dario Akaberi, Johan Lennerstrand, Åke Lundkvist
The RactIP server was used to predict long-range RNA-RNA interactions. The 5’ UTR and 3’ UTR sequences were given as input in separate fields. The server predicted the following interactions: 5’ GGAGAACAAG 3’ in the 5’ UTR with 5’ CUUGUUCUCC 3’ in the 3’ UTR. These sequences are inverted repeats. We found these sequences conserved in AFHV (114–123 in the 5’ UTR and 10695–10704 in the 3’ UTR; accession number: JF416957.1), KFDV (114–123 in the 5’ UTR and 10694–10703 in the 3’ UTR; accession number: JF416958.1 or HM055369.1) and TBEV (115–124 in the 5’ UTR and 11062–11071 in the 3’ UTR; accession number: NC_001672.1). This sequence has 50% GC content (total = 10 nucleotides). In all the three viruses, the repeat sequence in the 5’ UTR was found in one of the hairpin loops. In the 3’ UTR, on the other hand, the repeat sequence was predicted to be in one of the hairpin loops in KFDV (Figure 5, bottom), in one of the broken stems (bulges) in AFDV (Figure 5, bottom), and in one of the internal loops in TBEV (Figure 5, bottom).
A Simple Whole-Blood Polymerase Chain Reaction without DNA Extraction for Thalassemia Diagnosis
Published in Hemoglobin, 2018
Wibhasiri Srisuwan, Thanusak Tatu
Heme liberated from Hb has been proven to be a potent PCR inhibitor by direct inactivation of the thermostable DNA polymerase [6]. This is why the traditional PCR reaction was not possible when adding whole blood directly into the reaction. Al-Soud et al. [6] and Kreader [19] demonstrated that adding BSA or betaine to the reaction mixtures significantly reduced the inhibitory effect of blood. Thus, BSA and betaine were tested in this study, and betaine was found to be the best PCR facilitator in the presence of whole blood compared to BSA. Naturally, betaine is a polyglycine compound (N,N,N-trimethylglycine) [20]. It has been utilized to enhance amplification of GC-rich DNA template, due to its ability to reduce formation of secondary structures [21]. In whole-blood PCR, betaine may absorb heme, hence, removing the inhibitory effect of heme on DNA polymerase [6].
Novel strain of Pseudoruminococcus massiliensis possesses traits important in gut adaptation and host-microbe interactions
Published in Gut Microbes, 2022
Kaisa Hiippala, Imran Khan, Aki Ronkainen, Fredrik Boulund, Helena Vähä-Mäkilä, Maiju Suutarinen, Maike Seifert, Lars Engstrand, Reetta Satokari
The WGS and 16S rRNA sequence data helped in the species identification and taxonomical assignment resulting in the highest similarity to the type strain P. massiliensis Marseille-P3876 T (= CSUR P3876). The ORTHO ANI software15 resulted in average nucleotide sequence identity (ANI) of 97.98% between the genomes of the type strain of P. massiliensis and isolate E10-96H (Figure 1b). Similarly, 100% sequence identity was observed for 16S rRNA gene sequences of P. massiliensis E10-96H and P. massiliensis type strain (Figure 1c) and both showed phylogenetic proximity with R. bromii. The Type (Strain) Genome Server (TYGS) was also used to cluster species and subspecies. The Genome Blast Distance Phylogeny (GBDP) approach of TYGS also assigned isolate P. massiliensis E10-96H to the type strain of P. massiliensis. The G + C content variation within species at genome level was less than 1%16 (a score of 0.51) which also supported reliable identification.