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Using p35 gene primers of AcNPV, about 1 kb fragment was obtained by PCR from HaNPV DNA and was sequenced thereafter. It has a full reading frame encode 299 amino acids. Sharing identity of 94% in nucleotides and 84% in predict amino-acids with AcNPV, a apoptosis inhibiting gene was found. According to comparison of p35 genes in five kinds of baculovirus, it is found that AcNPV, TnNPV and BmNPV share the most intimate blood relation. HaNPV is near the above three species. LsNPV is little remote from them. This result reconfirmed those we have done with gp37 genes and vp39 genes. It is more accurate to use conserved gene for species division than that of the serological identification. [ABSTRACT FROM AUTHOR]

Lithium chloride facilitates the softening of cell walls resulting in a simple, quick (2 h) method for DNA extraction from the red seaweed Porphyra perforata J. Agardh. A 5-min treatment of tissues in LiCl at 55°C extracts DNA that is relatively free of the viscous polysaccharides and proteins that are usually coextracted in large amounts from cells walls and cytoplasm. This protocol does not require grinding of tissues, hydroxyapatite binding, cetyl trimethyl ammonium bromide treatments, enzymatic treatments, phenol extraction, or CsCl-gradient quality to be used as a template for polymerase chain reaction amplification. DNA extraction, LiCl, PCR amplification, Porphyra perforata, Rhodophyta [ABSTRACT FROM AUTHOR]

Individual merRTΔP regions were amplified from DNA directly isolated from soil and sediment samples using consensus primers derived from the conserved mer sequences of Tn501, Tn21 and pMER419. Soil and sediment samples were taken from four sites in the British Isles; one 'pristine' (SB) and three polluted (SO, SE, T2) with respect to mercury. The sizes of the PCR products amplified (= 1 kb) were consistent with their generation from mer determinants related to the archetypal elements found in Gram negative bacteria. Forty-five individual clones of sequences obtained from these four sites were isolated which hybridized (> 70% homology) to a merRTΔP probe from Tn501. The diversity of these amplified mer genes was analysed using Restriction Fragment Length Polymorphism (RFLP) profiling. Fourteen RFLP classes were distinguished, 12 of which proved to be novel and only two of which had been identified in an earlier study of 40 Gram negative mercury resistant bacteria cultured from the same four sites. UPGMA analysis was used to examine the relationships between the 22 classes of determinant identified. The T2 site, which has the longest history of mercury exposure, was found to have the greatest level of diversity in terms of numbers of classes of determinant, while the SO site, which had the highest mercury levels showed relatively low variation. Variation of mer genes within and between the sequences from cultivated bacteria and from total bacterial DNA shows clearly that analysing only sequences from cultivated organisms results in a gross underestimation of genetic variation. [ABSTRACT FROM AUTHOR]

Amyloid deposition in 11 inbred strains of mice (A/J, SJL/J, DDD, C57BL/6J, B10.BR, C57BL/10, Bl0A/SgSn, C3H/HeMs, B10A(5R), DBA/2 and C57BL/6Cr5/c) was studied using the peroxidase antiperoxidase (PAP) method and antisera against AS and murine protein AA. Among the 170 mice examined, in 77 (45.3%) from the nine strains other than C3H/ HeMs and DBA/2, there was evidence of spontaneous amyloid deposits in routine histological sections. Immunohistochemical studies using 54 mice with amyloid deposition, demonstrated AS deposition in 45 mice (83.3%) in all nine strains, although the incidence and intensity of the deposition differed somewhat between strains. SJL/J and A/J had AS deposits from the age of 8 months and the incidence increased with advancing age. In the other seven strains, AS was first deposited at an older age than in the SJL/J and A/J strains. In A/J, C57BL/6J, C57BL/10, B10.BR, B10A(5R) and C57BL/6Cr5/c, protein AA often coexisted with AS The distribution pattern of the AS deposits was similar to that observed among the SAM strains. Thus, AS is an ubiquitously distributed senile amyloid protein in the mouse. Determination of the molecular type of apoA-II, a serum precursor of AS, among all 11 strains using the polymerase chain reaction (PCR) revealed the SAM-P/1 type apoA-II variant in SJL/J and A/J strains with a high susceptibility to AS deposition. We concluded from this study that amino acid substitution in precursor apoA-II may be responsible for the early onset and severe amyloid deposition in the mouse. [ABSTRACT FROM AUTHOR]

A method for preparing DNA for PCR has been adapted from the forensic work of Walsh et al. (Biotechniques 10:506-513) for use with Chlamydomonas reinhardtii and Arabidopsis thaliana. The method consists of a short incubation of cells or tissue in ethanol, followed by addition of Chelex-100 and heat treatment. Following centrifugation, the supernatant is added directly to the PCR reaction; for Chlamydomonas, amplification product is visible over a range of four orders of magnitude of starting cells. Using this method, DNA suitable for PCR template can also be obtained from Arabidopsis leaf tissue without grinding, organic extraction or precipitation steps. This method may prove to be useful for other plant and algal species. [ABSTRACT FROM AUTHOR]

A survey of 480 10-mer primers for RAPD markers revealed general consistency in primer amplification strength among the flowering plant genera Datisca, Helianthus and Yucca. Six characteristics of primer base sequences were analyzed: total (G+C) content; the amounts of G, A, C, and T taken separately; and the (G+C) content in the last four bases of the 3′ end. Of these, total (G+C) content showed the most value in predicting primer amplification strength. Since the consistency of amplification strength is true only globally, there are still many primers showing high variation in amplification strength among genera, most probably due to DNA sequence differences, but perhaps also resulting from experimental artifact. Nevertheless, we suggest that this survey be used as a rough guide for prioritizing primer deployment in RAPD studies involving plants in the hope of improving efficiency during the search for adequate levels of polymorphism, with the understanding that taxon-specific differences in primer amplification strength are bound to occur. [ABSTRACT FROM AUTHOR]

A procedure to test for DNA methylation at sites recognized by methylation-sensitive restriction endonucleases is described. The procedure is based on the assumption that the polymerase chain reaction (PCR) will amplify sequences between two primers only if the target DNA is intact after digestion. A carrot ( Daucus carota) cell line that is heterozygous for two sequenced alleles of Dc8, a gene which is expressed during the later stages of embryogenesis provided an ideal source of DNA for developing and testing protocols. The promoters of the two alleles differs significantly in length between two sites used for primers, and only one promoter has a GATC ( Sau 3A1 or Mbo I) site. This allowed development of a protocol where only the sequence lacking the GATC site was amplified to detectable levels following digestion of DNA with Mbo I which is insensitive to symmetric methylation withC orC. [ABSTRACT FROM AUTHOR]

We describe here methods and strategies for amplifying and sequencing the genes encoding the small subunits (16/18S) of nuclear and chloroplast ribosomal DNA (rDNA) from total plant DNA. These methods were developed in response to technical difficulties we encountered in our molecular systematic work with members of various plant families. These protocols have proved useful when the amount of tissue available for study is limited and when the tissues have high concentrations of undesirable secondary metabolites which are often co-isolated with nucleic acids. [ABSTRACT FROM AUTHOR]

We describe a general method for the preparation of λZAP II cDNA libraries from very small amounts (<50 mg) of plant tissue. We have achieved this by combining an efficient method for RNA extraction with a modified PCR protocol for the synthesis and amplification of cDNA. Using this protocol we have found it possible to generate cDNA libraries containing more than 10 clones from as little as 1 μg of total RNA. [ABSTRACT FROM AUTHOR]

The human Rh blood-group system is encoded by two homologous genes, RhD and RhCE. The RH genes in gorillas and chimpanzees were investigated to delineate the phylogeny of the human RH genes. Southern blot analysis with an exon 7-specific probe suggested that gorillas have more than two RH genes, as has recently been reported for chimpanzees. Exon 7 was well conserved between humans, gorillas, and chimpanzees, although the exon 7 nucleotide sequences from gorillas were more similar to the human D gene, whereas the nucleotide sequences of this exon in chimpanzees were more similar to the human CE gene. The intron between exon 4 and exon 5 is polymorphic and can be used to distinguish the human D gene from the CE gene. Nucleotide sequencing revealed that the basis for the intron polymorphism is an Alu element in CE which is not present in the D gene. Examination of gorilla and chimpanzee genomic DNA for this intron polymorphism demonstrated that the D intron was present in all the chimpanzees and in all but one gorilla. The CE intron was found in three of six gorillas, but in none of the seven chimpanzees. Sequence data suggested that the Alu element might have previously been present in the chimpanzee RH genes but was eliminated by excision or recombination. Conservation of the RhD gene was also apparent from the complete identity between the 3′-noncoding region of the human D cDNA and a gorilla genomic clone, including an Alu element which is present in both species. The data suggest that at least two RH genes were present in a common ancestor of humans, chimpanzees, and gorillas, and that additional RH gene duplication has taken place in gorillas and chimpanzees. The RhCE gene appears to have diverged more than RhD among primates. In addition, the RhD gene deletion associated with the Rh-negative phenotype in humans seems to have occurred after speciation. [ABSTRACT FROM AUTHOR]

The human Rh blood group locus consists of two structurally related genes ( D and CcEe) in Rh-positive haplotypes but a single gene ( CcEe) in Rh-negative haplotypes. The genome of rhesus monkeys ( Macaca mulatta), while not expressing any of the human Rh D, C, c, E, or e specificities, carries a Rh-like locus strongly related to the human Rh locus. Southern blot analysis suggested the presence of only one Rh-like gene with an additional truncated fragment corresponding to the 5′ region. RNA preparations from M. mulatta bone marrow cells contained Rh-like species of 1.7 kb. Two allelic Rh-like transcripts were amplified by PCR and sequenced. The predicted translation product of the first transcript was a 417-amino-acid protein closely similar to the human Rh counterpart. The predicted product of the second transcript consisted of a 361-amino-acid polypeptide truncated in the NH terminal region and differing from the former by a few substitutions. The macaque Rh-like protein sequences differed from those of human D and Cc/Ee polypeptides by 22-25%, whereas the degree of identity between the human proteins was 91.5%. Implications of these results in the analysis of the evolutionary pathway of the Rh locus are discussed. [ABSTRACT FROM AUTHOR]

The nuclear capsid protein gene (vp39) of Bombyx mori nuclear polyhedrosis virus (BmNPV) was amplified successfully by PCR technique and inserted into pGEM 3zf(+). The 5′ and 3′ terminal area of the amplified vp39 gene were sequenced with silver-staining dideoxy method. Bmvp39 gene was sub-cloned into the expression vector pRSET-A, and transformed into E. coli BL21. This gene was highly expressed by IPTG induction. SDS-PAGE analysis showed that the expressed protein is about 38 kd, and the expressed amount reached maxium in 4 h with IPTG induction. [ABSTRACT FROM AUTHOR]

cDNA fragments from ten different protein kinases expressed in Avena sativa aleurone cells were amplified from mRNA by RT-PCR with degenerate primers. These could be classified into five groups: Aspk1-3 showed homology to the Snf1-related protein kinases, Aspk4-5 to a wheat ABA up-regulated protein kinase, Aspk6-8 to the Ca-dependent, calmodulin-independent protein kinase family, Aspk9 encoded a MAP kinase and Aspk10 was closely related to a novel Arabidopsis ribosomal protein kinase. GA caused a rapid increase in transcripts hybridising to Aspk10, while inhibiting the dramatic accumulation of transcripts hybridising to Aspk9 that occurred in the absence of GA. [ABSTRACT FROM AUTHOR]

The small subunit ribosomal DNA for a woody actinorhizal, Alnus glutinosa, was isolated by the PCR method. Amplification products were cloned into the Bluescript SK vector. Full sequence, 1698 bp, was obtained with NS1 to NS8 primers. Sequence alignments were made by UWGCG sequence data analysis computer programs. 18S rDNA sequence of A. glutinosa was compared to analogous segments of four other angiosperms, tomato, rice, maize and soybean. Sequence homologies are discussed and application for the technique is suggested. [ABSTRACT FROM AUTHOR]

A simple technique for direct sequencing of PCR-amplified templates without purification of the PCR reaction product is presented. This method does not require an additional synthesis step after template amplification, and can generate sequence information form as little as 0.1 fmol of unpurified template. [ABSTRACT FROM AUTHOR]

The advent of molecular genetics has had a significant impact on carrier detection and prenatal diagnosis in the haemophilias. Where phenotypic analysis can only identify with varying degrees of probability up to 90% of carriers and can only be used in prenatal diagnosis when fetal blood is obtained (18-20 weeks gestation), genotypic analysis gives a greater than 99% certainty of carrier status (when informative) and allows for prenatal diagnosis at 8-12 weeks utilising DNA obtained by chorionic villus sampling. Furthermore, the introduction of the technique of polymerase chain reaction (PCR) amplification of DNA into genotypic analysis either for the detection of the disease-causing defect itself, or to detect an intragenic informative polymorphism, has significantly increased the ease by which these procedures can be introduced into the laboratory. PCR based family studies in haemophilia will become increasingly available in both developed and developing countries. While in the former detection of the defect itself will become available, particularly for haemophilia B, in other countries simple PCR based DNA polymorphism analysis will become the mainstay of effective, practical haemophilia genetics. [ABSTRACT FROM AUTHOR]

The major simple sequence repeats present in the Arabidopsis genome were identified by Southern hybridizations with 49 oligonucleotide probes matching all the possible combinations of motifs up to 4 nucleotides long. The method used allowed us to perform all the hybridizations under the same temperature conditions. A good correlation was observed with the data obtained from database analysis, indicating that the method can be useful for identifying the major classes of microsatellite loci in species for which few or no sequence data are available. AG/CT, AAG/CTT, ATG/CAT and GTG/CAC are the major motifs present in the Arabidopsis genome that can be used as convenient probes to isolate microsatellite loci by screening libraries. AAG/CTT is the more frequent of these motifs, and its relative frequency in Arabidopsis is much higher than averagely found in the plant kingdom. About 8% of the cDNA clones from an immature silique library contains AG/CT, AAG/CTT or ATG/CAT microsatellite loci. Several microsatellite loci were isolated by screening genomic and cDNA libraries. Twenty-six tri-nucleotide loci were PCR amplified from four different ecotypes, and polymorphism was observed for 12 of them; 10 loci showing two alleles and 2 loci showing three alleles. [ABSTRACT FROM AUTHOR]

We describe a set of novel primers for successful amplification of the complete mitochondrial cytochrome b (cyt b) gene of ungulate species. DNA extracted from non-invasively obtained and decomposed samples is found to be degraded and inappropriate for amplification of the complete gene (more than 1 kb) in single PCR amplification. We developed a series of six ungulate-specific conserved primers for the cyt b gene. These primers, in various combinations, amplify 366-1,266 bp fragments. We also used them in multiplex PCR reaction to assess the maximum possible size of the PCR amplification product. [ABSTRACT FROM AUTHOR]

Amplification of high GC content genes by PCR is a major challenge during the creation of recombinant GC-rich DNA constructs. This may be due to the difficulty in DNA denaturation or the possibility of forming secondary structures from DNA templates. Tools have been described to address the technical problems associated with the amplification of shorter sequences (<1000 bp). However, obstacles of synthesizing larger-sized GC-rich sequences by PCR continue to exist. This study aims to investigate the amplification of long and high GC content genes by PCR from the Mycobacterium bovis , a genome with GC content >60%, in comparison to amplifying a gene from the Listeria monocytogenes genome, a genome with a 37.8% GC content. Three PCR protocols were designed and experimented at various conditions with two M. bovis genes, Mb0129 , a large gene of 1794 bp with 77.5% GC content, mpb83 , a smaller gene of 663 bp in length with moderate GC content of 63%, together with LMHCC_RS00060 , a large L. monocytogenes gene of 1617 bp with a lower GC content of 41.5%. The result demonstrated the superiority of the 2-step PCR protocol over other protocols in PCR amplification of Mb0129 when specific high fidelity DNA polymerases were used in the presence of an enhancer. The study highlighted the importance of manipulating the cycling conditions to perform the annealing and extension steps at higher temperatures while adjusting the ramp speed at a lower speed for a successful PCR amplification of a large GC-rich DNA template. A final PCR protocol was developed and enabled the amplification of 51 GC-rich targets. This can be a valuable tool for the amplification of long GC-rich DNA sequences for various downstream applications. • Lengthy GC rich DNA sequences pose an obstacle to successful PCR amplification. • Amplification of GC-rich targets from M. bovis for cloning is challenging. • Two step PCR using a slow thermal cycler for amplifying lengthy GC-rich targets. • A final protocol to amplify lengthy GC-rich targets without individual optimization. [ABSTRACT FROM AUTHOR]

• New molecular markers are available to detect DNA of grape berry moth from bat faeces. • Bat activity increases when grape berry moths are present in vineyards. • Eleven bat species are identified as predators of grape berry moths. • Bats are delivering pest control services in French vineyards. Faced with current health and environmental challenges, viticulture is directly concerned with the need to reduce pesticide use. Natural pest control services provided by bats have been demonstrated in other crops and is regularly mentioned as a way to reduce pesticide use. However, the trophic link between bats and grape pests as well as the effect of pest presence on bat activities remain largely unknown. To investigate the functional role of bats in vineyard landscapes, we used two independent approaches. We monitored the activities of bats and of the European grapevine moth (Lobesia botrana) in 23 vineyards located in the Bordeaux region (France). In parallel, we developed DNA primers to examine bat faeces from two regions, Bordeaux and Burgundy, for the presence of the three main species of grapevine moths. Our results demonstrate that bats significantly increase their hunting activity when European grapevine moths are present in vineyards. In addition, our molecular analysis of the faeces provides robust evidence that at least 10 species of bats predate the three grapevine moth species. Our results therefore suggest that bats can be natural enemies of grape pests in vineyard landscapes. Further research is now needed to investigate the consequences of predation of pests by bats on crop production as well as the effect of some management options at both the local and landscape scale to increase the level of pest control services provided by bats. [ABSTRACT FROM AUTHOR]

The direct amplification by PCR of the DNA in tissue samples of eggs and neonate larvae of Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae) was accomplished, and it is a new alternative for DNA amplification from fall armyworm samples. This method is simple, fast, economic, and accelerates studies on this polyphagous pest. [ABSTRACT FROM AUTHOR]

Purpose: Detection of the virulence gene is a key component in determining the pathogenicity of any isolates, because these genes act multi-functionally and multi-factorially. A gyrase specific gene primer, in combination with the PCR technology, allows the precise detection of the DNA gyrase subunit B2 gene (gyr-B-2) from different virulent microorganisms. In the present study, forward and reverse primers with lengths of 20bp and 21bp were used for the detection of the gyr-B-2 genes in the clinical isolates of the Klebsiella sp. which were collected from patients who were suffering from pneumonia and urinary tract infections (UTIs).Materials and Methods: A total of 14 isolates viz., K1, K2, K3, K4, K5, K6, K7, K8, K9, K10, K11, K12, K13 and K14 were isolated from 3 different private medical colleges of Sylhet city. Results: The gyr-B-2 gene which was amplified in 12 isolates viz., K1, K2, K3, K4, K5, K6, K7, K8, K10, K11, K12 and K14 gave the expected 411bp PCR product after its visualization under a gel documentation system in a 1.2% agarose gel. Conclusions: The present study was undertaken to detect the gyrB2 gene from Klebsiella sp, which will be helpful for further scientific studies. This PCR was outstanding in the detection of the gyb-B-2 gene in pneumonia and urinary tract infections in patients, which were caused by the Klebsiella species. [ABSTRACT FROM AUTHOR]

Chicken (Gallus gallus), cattle (Bos taurus), goat (Capra hircus), pig (Sus scrofa domestica) and wild boar (Sus scrofa linneus) raw meats were examined using PCR amplification of mitchondrial DNA. Three oligoprimers were used to amplified mitochondria DNA (mtDNA) of cytochrome b (two types of primers) and mitochondrial 12s rRNA (mt-12s rDNA) (one type of primer) gene for vertebrate-specific. The amplification PCR products of 359 bp and 531 bp were successfully amplified from the cyt b gene of pig and wild boar meats using two type of mtDNA. None of the band was observed for chicken, cattle and goat. While, the amplification product of all meats using mt-12S rDNA gene were successfully produced a single band with molecular size of 456 bp, which were as expected due to all animals were veterbrate-speciic. Thus, this primer could not be used to detect the pig DNA in raw meat samples. In the present work, the PCR amplification of mtDNA of cytochrome b has been shown as a suitable tool for rapid detection of pig DNA in foods. [ABSTRACT FROM AUTHOR]

Abstract: A procedure for the isolation of target PCR bands from multiplex PCR products using the E-Gel® iBase Power System, an E-Gel® Safe Imager™ Real-Time Transilluminator, and E-Gel® CloneWell 0.8% SYBR Safe™ agarose cassettes is presented. The collected isolates are suitable for direct use in sequencing reactions without need of further purification. Reductions in time spent per sample and cost per sequence produced compared to traditional “band-cutting” methods are demonstrated. An added benefit is that the new procedure does not require exposure to ethidium bromide or ultraviolet radiation. [Copyright &y& Elsevier]

Abstract: Here we report an improved, reproducible, simple, rapid, and cost-effective PCR-based DNA synthesis method using short (25–40 bp) overlapping oligodeoxyribonucleotides (oligos). The method involves two steps; (1) assembly of multiple/overlapping oligos by PCR to generate the template DNA and (2) amplification of the template DNA sequence with the two outermost oligos as primers. We have tested this method by synthesizing approximately 35 genes ranging in size between 300 bp and 1700 bp and G+C content from moderate (30%) to high (65%). In addition, we used the method to introduce 29 mutations simultaneously into a single gene. Key to the success of this method is the use of optimized oligo concentrations and the type of DNA polymerase used. This simplified and highly reproducible method is expected to be beneficial for the synthesis of a wide variety of genes. [Copyright &y& Elsevier]

Any forms of valorization of microorganisms would require accurate identity recognition to ensure repeatability, reproducibility and quality assurance. This study aimed to evaluate the effectiveness of different primers for identifying cultured eukaryotic microalgae using a simple 18S rDNA approach. A total of 34 isolated microalgae and one culture collection were utilized in the search for an effective molecular identification method for microalgae. Ammonium formate was applied to marine microalgae prior to DNA extraction. The microalgal DNA was extracted using a commercial kit and subjected directly to PCR amplification using four different published 18S rDNA primers. The DNA sequences were analysed using Basic Local Alignment Search Tool (BLAST) and phylogenetic trees to determine the microalgae identity. The identity was further validated with conventional morphological taxonomic identification, and the relationship of microalgal morphology and genetic materials was also determined. The microalgal DNA was successfully amplified, including marine species without prior cleaning. In addition, the ss5 + ss3 primer pair was found to be an ideal primer set among the tested primers for identifying microalgae. Overall, molecular identification showed relative matching with morphological identification (82.86%). This study is important because it serves as a platform to develop a standardized eukaryotic microalgae identification method. In addition, this method could help to ease the eukaryotic microalgae identification process and enrich the current reference databases such as GenBank. • Effectiveness of different published primers was tested on 35 cultured microalgae. • Marine microalgae do not require desalting process prior to PCR amplification. • Same molecular identification approach could apply to freshwater and marine microalgae. • ss5 + ss3 primer pair showed the highest universality in identifying microalgae. • 82.86% matching between molecular and morphological identification at the genus level. [ABSTRACT FROM AUTHOR]

Five simplified DNA preparation procedures for polymerase chain reaction (PCR) amplification were tested for detection of phytoplasmas from infected herbaceous and woody plants. Thin freehand cross-sections made from infected plant tissues and stored in acetone were used as sources for DNA preparation. The tissue sections were treated by: 1) grinding in sodium hydroxide; 2) sonicating in water; 3) microwaving in water; 4) boiling in sodium hydroxide; or 5) placing directly in PCR tube. PCR amplification was performed with a universal phytoplasma-specific primer pair in a reaction buffer containing 0.5% (v/v) Triton X-100, 1.5 mM magnesium chloride, and 10 mM Tris-HCl. All five procedures provided phytoplasmal template DNA for successful PCR amplification from infected herbaceous plants {periwinkle [Catharanthus roseus (L.) G. Don (periwinkle)], carrot (Daucus carota L.), maize (Zea mays L.)}, while the grinding, microwaving, and boiling procedures also allowed positive amplification from a woody plant [green ash (Fraxinus pennsylvanica Marsh.)]. The quality of the resulting DNA was adequate for subsequent identification of the aster yellows and ash yellows phytoplasmas through nested-PCR using phytoplasma group-specific primer pairs. These methods provide remarkable savings in labor and materials, making disease testing and indexing of plant materials much more attractive. [ABSTRACT FROM AUTHOR]

In this chapter, we provide a methodological description of the process to perform gastrointestinal (GIT) microbiota profiling on human stool samples. The process includes: (i) collection of feces, (ii) isolation of DNA from fecal community bacteria, (iii) selection of both 16S rDNA sequencing target and next-generation sequencing platform, and (iv) analysis and interpretation of sequence data. The process culminates into a comprehensive report on the GIT microbiota composition and structure that may translate into clinically actionable results.