A genetical approach to the physical mapping of chloroplast genes in Chlamydomonas

1984 ◽  
Vol 62 (4) ◽  
pp. 225-229 ◽  
Author(s):  
Claude Lemieux ◽  
Monique Turmel ◽  
Verner L. Seligy ◽  
Robert W. Lee
Keyword(s):  
16S Rrna ◽  
Chloroplast Dna ◽  
Genetic Markers ◽  
Streptomycin Resistance ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Chloroplast Genes ◽  
Restriction Fragments ◽  
Dna Restriction Fragments ◽  
Dna Restriction

Among the 100 potential chloroplast genes encoding polypeptides in higher plants, only 8 have been identified and mapped so far. Physical mapping of additional polypeptide genes for which there is no molecular probe can be facilitated by taking advantage of chloroplast genetics. We have demonstrated that it is possible to use a combination of genetical and molecular approaches to map directly a gene locus on the chloroplast genome of the interfertile green algae Chlamydomonas eugametos and Chlamydomonas moewusii. Because of the extensive recombination of chloroplast DNA and the biparental inheritance of chloroplast genetic markers detected in C. eugametos – C. moewusii hybrids, the inheritance of such genetic markers can be correlated with that of certain chloroplast DNA restriction fragments characteristic of one or the other parent. In 41 hybrids, a strict correlation was noted between the inheritance of a streptomycin resistance marker and that of chloroplast DNA restriction fragments encoding the two 16S rRNA genes. These results, therefore, support the view that the streptomycin resistance locus resides in each of the two 16S rRNA genes.

The chloroplast genome of Pinussylvestris; physical map and localization of chloroplast genes

1993 ◽  
Vol 23 (2) ◽  
pp. 234-238 ◽  
Author(s):  
B. Karpinska ◽  
S. Karpinski
Keyword(s):  
16S Rrna ◽  
Chloroplast Dna ◽  
Gene Order ◽  
Restriction Site ◽  
Physical Map ◽  
Genetic Maps ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Chloroplast Genes ◽  
Restriction Fragments

A physical map of Pinussylvestris L. chloroplast DNA for three restriction endonucleases (PstI, SacI, and KpnI) has been prepared by hybridization of isolated or cloned restriction fragments. Pinussylvestris chloroplast DNA is about 120 kilobases. Ten chloroplast genes have been localized by hybridization with heterologous chloroplast DNA probes. The genome contains single copies of the 23S and 16S rRNA genes and lacks any large repeated sequences. The restriction site arrangement and gene order have been compared with other known chloroplast DNA genetic maps of pine species.

scholarly journals Formation of Pseudo-Terminal Restriction Fragments, a PCR-Related Bias Affecting Terminal Restriction Fragment Length Polymorphism Analysis of Microbial Community Structure

2003 ◽  
Vol 69 (5) ◽  
pp. 2555-2562 ◽  
Author(s):  
Markus Egert ◽  
Michael W. Friedrich
Keyword(s):  
Restriction Fragment Length Polymorphism ◽  
16S Rrna ◽  
Length Polymorphism ◽  
Fragment Length Polymorphism ◽  
Rflp Analysis ◽  
Restriction Enzymes ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Restriction Fragments ◽  
Restriction Fragment Length

ABSTRACT Terminal restriction fragment length polymorphism (T-RFLP) analysis of PCR-amplified genes is a widely used fingerprinting technique in molecular microbial ecology. In this study, we show that besides expected terminal restriction fragments (T-RFs), additional secondary T-RFs occur in T-RFLP analysis of amplicons from cloned 16S rRNA genes at high frequency. A total of 50% of 109 bacterial and 78% of 68 archaeal clones from the guts of cetoniid beetle larvae, using MspI and AluI as restriction enzymes, respectively, were affected by the presence of these additional T-RFs. These peaks were called “pseudo-T-RFs” since they can be detected as terminal fluorescently labeled fragments in T-RFLP analysis but do not represent the primary terminal restriction site as indicated by sequence data analysis. Pseudo-T-RFs were also identified in T-RFLP profiles of pure culture and environmental DNA extracts. Digestion of amplicons with the single-strand-specific mung bean nuclease prior to T-RFLP analysis completely eliminated pseudo-T-RFs. This clearly indicates that single-stranded amplicons are the reason for the formation of pseudo-T-RFs, most probably because single-stranded restriction sites cannot be cleaved by restriction enzymes. The strong dependence of pseudo-T-RF formation on the number of cycles used in PCR indicates that (partly) single-stranded amplicons can be formed during amplification of 16S rRNA genes. In a model, we explain how transiently formed secondary structures of single-stranded amplicons may render single-stranded amplicons accessible to restriction enzymes. The occurrence of pseudo-T-RFs has consequences for the interpretation of T-RFLP profiles from environmental samples, since pseudo-T-RFs may lead to an overestimation of microbial diversity. Therefore, it is advisable to establish 16S rRNA gene sequence clone libraries in parallel with T-RFLP analysis from the same sample and to check clones for their in vitro digestion T-RF pattern to facilitate the detection of pseudo-T-RFs.

scholarly journals Increase in Terminal Restriction Fragments of Bacteroidetes-Derived 16S rRNA Genes after Administration of Short-Chain Fructooligosaccharides

2006 ◽  
Vol 72 (9) ◽  
pp. 6271-6276 ◽  
Author(s):  
Yusuke Nakanishi ◽  
Koichiro Murashima ◽  
Hiroki Ohara ◽  
Takahisa Suzuki ◽  
Hidenori Hayashi ◽  
...  
Keyword(s):  
16S Rrna ◽  
Intestinal Microbiota ◽  
Fragment Length Polymorphism ◽  
Rflp Analysis ◽  
Intestinal Bacteria ◽  
Short Chain ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Restriction Fragments ◽  
Culture Independent

ABSTRACT It is well known that short chain fructooligosaccharides (scFOS) modify intestinal microbiota in animals as well as in humans. Since most murine intestinal bacteria are still uncultured, it is difficult for a culturing method to detect changes in intestinal microbiota after scFOS administration in a mouse model. In this study, we sought markers of positive change in murine intestinal microbiota after scFOS administration using terminal restriction fragment length polymorphism (T-RFLP) analysis, which is a culture-independent method. The T-RFLP profiles showed that six terminal restriction fragments (T-RFs) were significantly increased after scFOS administration. Phylogenetic analysis of the 16S rRNA partial gene sequences of murine fecal bacteria suggested that four of six T-RFs that increased after scFOS administration were derived from the 16S rRNA genes of the class Bacteroidetes. Preliminary quantification of Bacteroidetes by real-time PCR suggests that the 16S rRNA genes derived from Bacteroidetes were increased by scFOS administration. Therefore, the T-RFs derived from Bacteroidetes are good markers of change of murine intestinal microbiota after scFOS administration.

Taxonomic patterns and inheritance of chloroplast DNA variation in a survey of Pinusechinata, Pinuselliottii, Pinuspalustris, and Pinustaeda

1992 ◽  
Vol 22 (5) ◽  
pp. 683-689 ◽  
Author(s):  
D.B. Wagner ◽  
W.L. Nance ◽  
C.D. Nelson ◽  
T. Li ◽  
R.N. Patel ◽  
...  
Keyword(s):  
Chloroplast Dna ◽  
Joint Analysis ◽  
Preliminary Screening ◽  
Dna Variation ◽  
Restriction Fragments ◽  
Two Samples ◽  
Dna Restriction Fragments ◽  
Chloroplast Dna Variation ◽  
Dna Restriction ◽  
Chloroplast Dna Markers

We used 20 heterologous probes (representing approximately 78% of the Pinuscontorta Dougl. chloroplast genome) and 16 endonucleases to examine chloroplast DNA restriction fragments from four Pinus L. species of the southeastern United States. Three variable markers, separated from each other by at least 20 kilobase pairs, were detected in a preliminary screening of two samples from each of the four southern pine species. Restriction fragments of these three markers were paternally inherited in controlled matings, as would be expected of chloroplast DNA genotypes of conifers. Eight chloroplast DNA haplotypes were identified by joint analysis of the three markers in a geographic survey of 215 individuals. The taxonomic distribution of these eight haplotypes placed the four southern pines in the following three groups: (i) Pinusechinata Mill.–Pinuspalustris Mill., (ii) Pinuselliottii Engelm., and (iii)Pinustaeda L. Little intraspecific variation was evident in the geographic survey, but one of the three chloroplast DNA markers was clearly polymorphic in P. elliottii.

Differentiation Of Clarias Batrachus, C. Gariepinus And Heteropneustes Fossilis By Pcr-Sequencing Of Mitochondrial 16s Rrna Gene

2015 ◽  
Vol 41 (1) ◽  
pp. 51-58
Author(s):  
Mohammad Shamimul Alam ◽  
Hawa Jahan ◽  
Rowshan Ara Begum ◽  
Reza M Shahjahan
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Fish Larva ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Valuable Insight ◽  
Multiple Sequence ◽  
Pcr Rflp ◽  
Alternative Means

Heteropneustesfossilis, Clariasbatrachus and C. gariepinus are three major catfishes ofecological and economic importance. Identification of these fish species becomes aproblem when the usual external morphological features of the fish are lost or removed,such as in canned fish. Also, newly hatched fish larva is often difficult to identify. PCRsequencingprovides accurate alternative means of identification of individuals at specieslevel. So, 16S rRNA genes of three locally collected catfishes were sequenced after PCRamplification and compared with the same gene sequences available from othergeographical regions. Multiple sequence alignment of the 16S rRNA gene fragments ofthe catfish species has revealed polymorphic sites which can be used to differentiate thesethree species from one another and will provide valuable insight in choosing appropriaterestriction enzymes for PCR-RFLP based identification in future. Asiat. Soc. Bangladesh, Sci. 41(1): 51-58, June 2015

scholarly journals Effects of Supplement of Marichromatium gracile YL28 on Water Quality and Microbial Structures in Shrimp Mariculture Ecosystems

Genes ◽  
2020 ◽  
Vol 12 (1) ◽  
pp. 40
Author(s):  
Liang Cui ◽  
Bitong Zhu ◽  
Xiaobo Zhang ◽  
Zhuhua Chan ◽  
Chungui Zhao ◽  
...  
Keyword(s):  
Water Quality ◽  
16S Rrna ◽  
Relative Abundance ◽  
Animal Health ◽  
Denitrifying Bacteria ◽  
Nitrite Reduction ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Sequencing Analysis ◽  
Shrimp Culture

The elevated NH3-N and NO2-N pollution problems in mariculture have raised concerns because they pose threats to animal health and coastal and offshore environments. Supplement of Marichromatium gracile YL28 (YL28) into polluted shrimp rearing water and sediment significantly decreased ammonia and nitrite concentrations, showing that YL28 functioned as a novel safe marine probiotic in the shrimp culture industry. The diversity of aquatic bacteria in the shrimp mariculture ecosystems was studied by sequencing the V4 region of 16S rRNA genes, with respect to additions of YL28 at the low and high concentrations. It was revealed by 16S rRNA sequencing analysis that Proteobacteria, Planctomycete and Bacteroidetes dominated the community (>80% of operational taxonomic units (OTUs)). Up to 41.6% of the predominant bacterial members were placed in the classes Gammaproteobacteria (14%), Deltaproteobacteria (14%), Planctomycetacia (8%) and Alphaproteobacteria (5.6%) while 40% of OTUs belonged to unclassified ones or others, indicating that the considerable bacterial populations were novel in our shrimp mariculture. Bacterial communities were similar between YL28 supplements and control groups (without addition of YL28) revealed by the β-diversity using PCoA, demonstrating that the additions of YL28 did not disturb the microbiota in shrimp mariculture ecosystems. Instead, the addition of YL28 increased the relative abundance of ammonia-oxidizing and denitrifying bacteria. The quantitative PCR analysis further showed that key genes including nifH and amoA involved in nitrification and nitrate or nitrite reduction significantly increased with YL28 supplementation (p < 0.05). The supplement of YL28 decreased the relative abundance of potential pathogen Vibrio. Together, our studies showed that supplement of YL28 improved the water quality by increasing the relative abundance of ammonia-oxidizing and denitrifying bacteria while the microbial community structure persisted in shrimp mariculture ecosystems.

scholarly journals Ultra-accurate microbial amplicon sequencing with synthetic long reads

Microbiome ◽  
2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Benjamin J. Callahan ◽  
Dmitry Grinevich ◽  
Siddhartha Thakur ◽  
Michael A. Balamotis ◽  
Tuval Ben Yehezkel
Keyword(s):  
Microbial Community ◽  
16S Rrna ◽  
Amplicon Sequencing ◽  
Species Level ◽  
Full Length ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Strain Identification ◽  
Long Reads ◽  
Long Read

Abstract Background Out of the many pathogenic bacterial species that are known, only a fraction are readily identifiable directly from a complex microbial community using standard next generation DNA sequencing. Long-read sequencing offers the potential to identify a wider range of species and to differentiate between strains within a species, but attaining sufficient accuracy in complex metagenomes remains a challenge. Methods Here, we describe and analytically validate LoopSeq, a commercially available synthetic long-read (SLR) sequencing technology that generates highly accurate long reads from standard short reads. Results LoopSeq reads are sufficiently long and accurate to identify microbial genes and species directly from complex samples. LoopSeq perfectly recovered the full diversity of 16S rRNA genes from known strains in a synthetic microbial community. Full-length LoopSeq reads had a per-base error rate of 0.005%, which exceeds the accuracy reported for other long-read sequencing technologies. 18S-ITS and genomic sequencing of fungal and bacterial isolates confirmed that LoopSeq sequencing maintains that accuracy for reads up to 6 kb in length. LoopSeq full-length 16S rRNA reads could accurately classify organisms down to the species level in rinsate from retail meat samples, and could differentiate strains within species identified by the CDC as potential foodborne pathogens. Conclusions The order-of-magnitude improvement in length and accuracy over standard Illumina amplicon sequencing achieved with LoopSeq enables accurate species-level and strain identification from complex- to low-biomass microbiome samples. The ability to generate accurate and long microbiome sequencing reads using standard short read sequencers will accelerate the building of quality microbial sequence databases and removes a significant hurdle on the path to precision microbial genomics.

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