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Molecular Biology

Single Genome Sequencing of Expressed and Proviral HIV-1 Envelope Glycoprotein 120 (gp120) and nef Genes

Authors: David J. Nolan
David J. NolanAffiliation 1: Bioinfoexperts, LLC, Thibodaux, Louisiana, USA
Affiliation 2: Department of Pathology, Immunology and Laboratory Medicine, Emerging Pathogens Institute, University of Florida, Gainesville, Florida, USA
Bio-protocol author page: a4654
, Susanna L. Lamers
Susanna L. LamersAffiliation: Bioinfoexperts, LLC, Thibodaux, Louisiana, USA
Bio-protocol author page: a4655
, Rebecca Rose
Rebecca RoseAffiliation: Bioinfoexperts, LLC, Thibodaux, Louisiana, USA
Bio-protocol author page: a4656
, James J. Dollar
James J. DollarAffiliation 1: Bioinfoexperts, LLC, Thibodaux, Louisiana, USA
Affiliation 2: Department of Pathology, Immunology and Laboratory Medicine, Emerging Pathogens Institute, University of Florida, Gainesville, Florida, USA
Bio-protocol author page: a4657
, Marco Salemi
Marco SalemiAffiliation: Department of Pathology, Immunology and Laboratory Medicine, Emerging Pathogens Institute, University of Florida, Gainesville, Florida, USA
Bio-protocol author page: a4658
 and Michael S. McGrath
Michael S. McGrathAffiliation 1: Departments of Laboratory Medicine, Pathology, and Medicine, University of California at San Francisco, San Francisco, California, USA
Affiliation 2: The AIDS and Cancer Specimen Resource, University of California at San Francisco, San Francisco, California, USA
For correspondence: MMcGrath@php.ucsf.edu
Bio-protocol author page: a4659
,  date: 6/20/2017, 180 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2334.

[Abstract] The current study provides detailed protocols utilized to amplify the complete HIV-1 gp120 and nef genes from single copies of expressed or integrated HIV present in fresh-frozen autopsy tissues of patients who died while on combined antiretroviral therapy (cART) with no detectable plasma viral load (pVL) at death (Lamers et al., 2016a and 2016b; Rose ...

Next-generation Sequencing of the DNA Virome from Fecal Samples

Authors: Cynthia L. Monaco
Cynthia L. MonacoAffiliation: Department of Medicine, Washington University School of Medicine, St. Louis, USA
For correspondence: cmonaco@wustl.edu
Bio-protocol author page: a4038
 and Douglas S. Kwon
Douglas S. KwonAffiliation 1: Ragon Institute of MGH, MIT, and Harvard, Cambridge, USA
Affiliation 2: Division of Infectious Diseases, Massachusetts General Hospital and Harvard Medical School, Boston, USA
Bio-protocol author page: a4039
,  date: 3/5/2017, 883 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.2159.

[Abstract] Herein we describe a detailed protocol for DNA virome analysis of low input human stool samples (Monaco et al., 2016). This protocol is divided into four main steps: 1) stool samples are pulverized to evenly distribute microbial matter; 2) stool is enriched for virus-like particles and DNA is extracted by phenol-chloroform; 3) purified DNA is multiple-strand ...

Generation of Mitochondrial-nuclear eXchange Mice via Pronuclear Transfer

Authors: Robert A. Kesterson
Robert A. KestersonAffiliation: Department of Genetics, University of Alabama, Birmingham, USA
For correspondence: kesterso@uab.edu
Bio-protocol author page: a3629
, Larry W. Johnson
Larry W. JohnsonAffiliation: Department of Genetics, University of Alabama, Birmingham, USA
Bio-protocol author page: a3630
, Laura J. Lambert
Laura J. LambertAffiliation: Department of Genetics, University of Alabama, Birmingham, USA
Bio-protocol author page: a3631
, Jay L. Vivian
Jay L. VivianAffiliation: Department of Pathology, University of Kansas Medical Center, Kansas City, USA
Bio-protocol author page: a3632
, Danny R. Welch
Danny R. WelchAffiliation: Department of Cancer Biology, University of Kansas Cancer Center, Kansas City, USA
For correspondence: dwelch@kumc.edu
Bio-protocol author page: a3633
 and Scott W. Ballinger
Scott W. BallingerAffiliation: Division of Molecular and Cellular Pathology, University of Alabama, Birmingham, USA
For correspondence: sballing@uab.edu
Bio-protocol author page: a3634
,  date: 10/20/2016, 1456 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1976.

[Abstract] The mitochondrial paradigm for common disease proposes that mitochondrial DNA (mtDNA) sequence variation can contribute to disease susceptibility and progression. To test this concept, we developed the Mitochondrial-nuclear eXchange (MNX) model, in which isolated embryonic pronuclei from one strain of species are implanted into an enucleated embryo ...

EST-SSR Analysis and Cross-species Transferability Study in Lavandula

Authors: Ayelign M. Adal
Ayelign M. AdalAffiliation: Department of Biology, University of British Columbia, Kelowna, Canada
Bio-protocol author page: a3398
, Zerihun A. Demissie
Zerihun A. DemissieAffiliation: National Research Council of Canada, Ottawa, Canada
Bio-protocol author page: a3399
 and Soheil S. Mahmoud
Soheil S. MahmoudAffiliation: Department of Biology, University of British Columbia, Kelowna, Canada
For correspondence: soheil.mahmoud@ubc.ca
Bio-protocol author page: a3400
,  date: 8/5/2016, 1524 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1891.

[Abstract] The genus Lavandula comprises of several economically important lavender species that are mainly cultivated worldwide for essential oil production. Identification of lavender species and their cultivars has been a huge bottleneck in lavender industries due to lack of appropriate identification mechanisms. Recent advances in modern technologies would ...

Identification of Natural Hybrids by SSR Markers in Mussaenda

Authors: Zhonglai Luo
Zhonglai LuoAffiliation: Key Laboratory of Plant Resources Conservation and Sustainable Utilization, South China Botanical Garden, the Chinese Academy of Sciences, China
Bio-protocol author page: a3261
, Tingting Duan
Tingting DuanAffiliation 1: Key Laboratory of Plant Resources Conservation and Sustainable Utilization, South China Botanical Garden, the Chinese Academy of Sciences, China
Affiliation 2: University of Chinese Academy of Sciences, China
Bio-protocol author page: a3262
, Shuai Yuan
Shuai YuanAffiliation 1: Key Laboratory of Plant Resources Conservation and Sustainable Utilization, South China Botanical Garden, the Chinese Academy of Sciences, China
Affiliation 2: University of Chinese Academy of Sciences, China
Bio-protocol author page: a3263
, Shi Chen
Shi ChenAffiliation: Beneficial Insects Institute, Fujian Agriculture and Forestry University, China
Bio-protocol author page: a3264
, Xiufeng Bai
Xiufeng BaiAffiliation: Department of Cell Biology, the Third Military Medical University, Chongqing, China
Bio-protocol author page: a3265
 and Dianxiang Zhang
Dianxiang ZhangAffiliation: Key Laboratory of Plant Resources Conservation and Sustainable Utilization, South China Botanical Garden, the Chinese Academy of Sciences, China
For correspondence: dx-zhang@scbg.ac.cn
Bio-protocol author page: a3266
,  date: 7/5/2016, 1499 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1853.

[Abstract] Detection of natural hybrids is of great significance for plant taxonomy, reproductive biology, and population genetic studies. Compared with methods depending on morphological characters, molecular markers provide reliable and much more accurate results. This protocol describes approaches employing microsatellite (SSR) markers to identify inter-specific ...

Genomic DNA Extraction and Genotyping of Dictyochloropsis Green Algae Strains

Authors: Francesco Dal Grande
Francesco Dal GrandeAffiliation: Biodiversity and Climate Research Centre (BiK-F), Senckenberg Gesellschaft fuer Naturforschung, Frankfurt am Main, Germany
For correspondence: francesco.dalgrande@senckenberg.de
Bio-protocol author page: a2413
, Carolina Cornejo
Carolina CornejoAffiliation: Biodiversity and Conservation Biology, Swiss Federal Research Institute WSL, Birmensdorf, Switzerland
Bio-protocol author page: a2328
, Christoph Scheidegger
Christoph ScheideggerAffiliation: Biodiversity and Conservation Biology, Swiss Federal Research Institute WSL, Birmensdorf, Switzerland
Bio-protocol author page: a2329
 and Andreas Beck
Andreas BeckAffiliation 1: Department of Lichenology and Bryology, Botanische Staatssammlung München, München, Germany
Affiliation 2: GeoBio-Center, Ludwig-Maximilians Universität München, München, Germany
For correspondence: beck@bsm.mwn.de
Bio-protocol author page: a2414
,  date: 8/5/2015, 2524 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1545.

[Abstract] Dictyochloropsis is an ecologically important genus of free-living and symbiotic green algae. Representatives of this genus are horizontally transmitted among several fungi of the family Lobariaceae, thus forming photobiont-mediated guilds. This protocol is suitable for extracting DNA from algal cultures and lichen samples and for genotyping seven ...

Pyrosequencing Approach for SNP Genotyping in Plants Using a M13 Biotinylated Primer

Authors: Cristina Silvar
Cristina SilvarAffiliation 1: Department of Animal and Plant Biology and Ecology, University of Coruña, Coruña, Spain
Affiliation 2: Institute for Resistance Research and Stress Tolerance, Julius Kuehn-Institute, Quedlinburg, Germany
Bio-protocol author page: a2212
, Dragan Perovic
Dragan PerovicAffiliation: Institute for Resistance Research and Stress Tolerance, Julius Kuehn-Institute, Quedlinburg, Germany
For correspondence: dragan.perovic@jki.bund.de
Bio-protocol author page: a2213
, Anna M Casas
Anna M CasasAffiliation: Department of Genetics and Plant Production, Aula Dei Experimental Station, CSIC, Zaragoza, Spain
Bio-protocol author page: a2215
, Ernesto Igartua
Ernesto IgartuaAffiliation: Department of Genetics and Plant Production, Aula Dei Experimental Station, CSIC, Zaragoza, Spain
Bio-protocol author page: a2216
 and Frank Ordon
Frank OrdonAffiliation: Institute for Resistance Research and Stress Tolerance, Julius Kuehn-Institute, Quedlinburg, Germany
Bio-protocol author page: a2214
,  date: 5/20/2015, 2640 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1473.

[Abstract] Single Nucleotide Polymorphisms (SNPs), which constitute single base-pair variations in the DNA sequence, are the most abundant molecular markers in plant and animal genomes. They are becoming the markers of choice for genotyping in all fields of molecular biology, as they are easily prone to automation and high throughput, for example through pyrosequencing. ...

Fluorescence-based CAPS Multiplex Genotyping on Capillary Electrophoresis Systems

Authors: Jelena Perovic*
Jelena PerovicAffiliation: Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Stadt Seeland, Germany
Bio-protocol author page: a2211
, Cristina Silvar*
Cristina SilvarAffiliation: Department of Animal and Plant Biology and Ecology, University of Coruña, Coruña, Spain
Bio-protocol author page: a2212
, Dragan Perovic
Dragan PerovicAffiliation: Institute for Resistance Research and Stress Tolerance, Julius Kuehn-Institute, Quedlinburg, Germany
For correspondence: dragan.perovic@jki.bund.de
Bio-protocol author page: a2213
, Nils Stein
Nils SteinAffiliation: Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Stadt Seeland, Germany
Bio-protocol author page: a1447
 and Frank Ordon
Frank OrdonAffiliation: Institute for Resistance Research and Stress Tolerance, Julius Kuehn-Institute, Quedlinburg, Germany
Bio-protocol author page: a2214
 (*contributed equally to this work),  date: 5/20/2015, 2465 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1472.

[Abstract] Recent advances in next-generation sequencing techniques allow the detection of a large number of SNPs and their use in a high throughput manner. However, Cleaved Amplified Polymorphic Sequences (CAPSs) still play a significant role as complement to other high throughput methods for SNP genotyping. Therefore, new methods focusing on the acceleration ...

Evolution of Escherichia coli to Macrophage Cell Line

Authors: Migla Miskinyte
Migla MiskinyteAffiliation: Evolutionary Biology Group, Instituto Gulbenkian de Ciência, Oeiras, Portugal
Bio-protocol author page: a1636
 and Isabel Gordo
Isabel GordoAffiliation: Evolutionary Biology Group, Instituto Gulbenkian de Ciência, Oeiras, Portugal
For correspondence: igordo@igc.gulbenkian.pt
Bio-protocol author page: a1637
,  date: 9/5/2014, 3110 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1227.

[Abstract] The genomes of species of Escherichia coli (E. coli) show an extraordinary amount of diversity, which include commensal strains and strains belonging to different pathovars. Many strains of E. coli, which can cause mild or severe pathologies in humans, have a commensal ancestor. Understanding the evolutionary changes that can lead to a transition from ...

Fitness Measurements of Evolved Esherichia coli

Authors: Migla Miskinyte
Migla MiskinyteAffiliation: Evolutionary Biology Group, Instituto Gulbenkian de Ciência, Oeiras, Portugal
Bio-protocol author page: a1636
 and Isabel Gordo
Isabel GordoAffiliation: Evolutionary Biology Group, Instituto Gulbenkian de Ciência, Oeiras, Portugal
For correspondence: igordo@igc.gulbenkian.pt
Bio-protocol author page: a1637
,  date: 9/5/2014, 3212 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1228.

[Abstract] Bacteria can adapt very rapidly to novel selective pressures. In the transition from commensalism to pathogenicity bacteria have to face and adapt to the host immune system. Specifically, the antagonistic interaction imposed by one of the first line of defense of innate immunity cells, macrophages, on commensal bacteria, such as Escherichia coli (E. ...
1 2 

[Bio101] Genotyping for Single Zebrafish (Fin Clip) or Zebrafish Embryo

Author: Lili Jing
Lili JingAffiliation: Department of Cell and Molecular Biology, University of Pennsylvania, Philadelphia, USA
For correspondence: lilijingcn@gmail.com
Bio-protocol author page: a38
,  date: 2/5/2012, 13733 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.182.

[Abstract] Zebrafish is increasingly used a genetic model organism in biomedical studies. This protocol provides a detailed procedure about the identification of the genotype of an adult zebrafish or a zebrafish embryo.
...

PCR-RFLP Genotyping of Point Mutations in Caenorhabditis elegans

Author: Peichuan Zhang
Peichuan ZhangAffiliation: Department of Biochemistry and Biophysics, University of California, San Francisco, USA
For correspondence: peichuan.zhang@ucsf.edu
Bio-protocol author page: a11
,  date: 3/20/2012, 12832 views, 3 Q&A
DOI: https://doi.org/10.21769/BioProtoc.128.

[Abstract] This protocol describes the basic principle of PCR/restriction digest genotyping of point mutations in worms, based on the principle of Restriction Fragment Length Polymorphism (RFLP) analysis. This type of genotyping is, particularly, useful when phenotypic analysis of animals carrying point mutations ...

A Quick, No Frills Approach to Mouse Genotyping

Author: Manuel E. Lopez
Manuel E. LopezAffiliation: Department of Developmental Biology, Stanford University, Stanford, USA
For correspondence: manduardo@gmail.com
Bio-protocol author page: a74
,  date: 8/5/2012, 12318 views, 1 Q&A
DOI: https://doi.org/10.21769/BioProtoc.244.

[Abstract] Mice are extremely powerful mammalian genetic model organisms for basic and medical research, but managing a colony of transgenic mice is time consuming and expensive, many times requiring the help of dedicated technicians. Slow and laborious genotyping procedures add to the hassle. Outsourcing is costly ...

[Bio101] Site-Directed Mutagenesis

Author: Lili Jing
Lili JingAffiliation: Department of Cell and Molecular Biology, University of Pennsylvania, Philadelphia, USA
For correspondence: lilijingcn@gmail.com
Bio-protocol author page: a38
,  date: 2/5/2011, 10885 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.29.

[Abstract] This protocol is for making nucleotide changes at specific loci in a large vector (>= 10 kb), and based on QuikChange II XL Site-Directed Mutagenesis Kit (stratagene)....

[Bio101] Genotyping Transgenic Zebrafish Using Genomic DNA Extracted from Clutch of Embryos (Originally by J. Lefebvre)

Author: Lili Jing
Lili JingAffiliation: Department of Cell and Molecular Biology, University of Pennsylvania, Philadelphia, USA
For correspondence: lilijingcn@gmail.com
Bio-protocol author page: a38
,  date: 2/5/2012, 8340 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.181.

[Abstract] Transgenic zebrafish are very useful genetic tools to study various biological processes. Identification the right transgene founder and the subsequent transgenic animals are always tedious and time consuming. This protocol provides a relatively rapid and easy method to identify the founder parent using ...

High-throughput Method for Determination of Seed Paternity by Microsatellite Markers

Authors: Samik Bhattacharya
Samik BhattacharyaAffiliation: Department of Molecular Ecology, Max Planck Institute for Chemical Ecology, Jena, Germany
Bio-protocol author page: a582
 and Ian T. Baldwin
Ian T. BaldwinAffiliation: Department of Molecular Ecology, Max Planck Institute for Chemical Ecology, Jena, Germany
For correspondence: baldwin@ice.mpg.de
Bio-protocol author page: a581
,  date: 4/20/2013, 5014 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.452.

[Abstract] In this protocol, determination of seed paternity by microsatellite markers in Nicotiana attenuata is described. However, this does not include a protocol for the novel marker selection/identification, but rather exploits the markers generated for a closely related species N. tabacum (Bindler et al., ...

Detection of Transposable Element Insertion Site Polymorphisms by Sequence-Specific Amplification Polymorphism (SSAP)

Authors: Véronique Sarilar
Véronique SarilarAffiliation: AgroParisTech, Unité Micalis - Équipe BimLip, Paris, France
For correspondence: veronique.sarilar@grignon.inra.fr
Bio-protocol author page: a1179
, Paulina Martinez Palacios
Paulina Martinez PalaciosAffiliation: AgroParisTech, Unité Micalis - Équipe BimLip, Paris, France
Bio-protocol author page: a1180
 and Karine Alix
Karine AlixAffiliation: AgroParisTech, Unité Micalis - Équipe BimLip, Paris, France
For correspondence: alix@moulon.inra.fr
Bio-protocol author page: a1181
,  date: 3/5/2014, 3379 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1054.

[Abstract] Transposable elements represent a major part of any eukaryotic genomes. Notably in plants they can account for more than 80% of the whole genomic sequence (such as in maize). Due to their mobility across the genome, they can act as mutagens but can also be considered as an important source of genetic ...

Fitness Measurements of Evolved Esherichia coli

Authors: Migla Miskinyte
Migla MiskinyteAffiliation: Evolutionary Biology Group, Instituto Gulbenkian de Ciência, Oeiras, Portugal
Bio-protocol author page: a1636
 and Isabel Gordo
Isabel GordoAffiliation: Evolutionary Biology Group, Instituto Gulbenkian de Ciência, Oeiras, Portugal
For correspondence: igordo@igc.gulbenkian.pt
Bio-protocol author page: a1637
,  date: 9/5/2014, 3212 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1228.

[Abstract] Bacteria can adapt very rapidly to novel selective pressures. In the transition from commensalism to pathogenicity bacteria have to face and adapt to the host immune system. Specifically, the antagonistic interaction imposed by one of the first line of defense of innate immunity cells, macrophages, ...

Evolution of Escherichia coli to Macrophage Cell Line

Authors: Migla Miskinyte
Migla MiskinyteAffiliation: Evolutionary Biology Group, Instituto Gulbenkian de Ciência, Oeiras, Portugal
Bio-protocol author page: a1636
 and Isabel Gordo
Isabel GordoAffiliation: Evolutionary Biology Group, Instituto Gulbenkian de Ciência, Oeiras, Portugal
For correspondence: igordo@igc.gulbenkian.pt
Bio-protocol author page: a1637
,  date: 9/5/2014, 3110 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1227.

[Abstract] The genomes of species of Escherichia coli (E. coli) show an extraordinary amount of diversity, which include commensal strains and strains belonging to different pathovars. Many strains of E. coli, which can cause mild or severe pathologies in humans, have a commensal ancestor. Understanding the evolutionary ...

Pyrosequencing Approach for SNP Genotyping in Plants Using a M13 Biotinylated Primer

Authors: Cristina Silvar
Cristina SilvarAffiliation 1: Department of Animal and Plant Biology and Ecology, University of Coruña, Coruña, Spain
Affiliation 2: Institute for Resistance Research and Stress Tolerance, Julius Kuehn-Institute, Quedlinburg, Germany
Bio-protocol author page: a2212
, Dragan Perovic
Dragan PerovicAffiliation: Institute for Resistance Research and Stress Tolerance, Julius Kuehn-Institute, Quedlinburg, Germany
For correspondence: dragan.perovic@jki.bund.de
Bio-protocol author page: a2213
, Anna M Casas
Anna M CasasAffiliation: Department of Genetics and Plant Production, Aula Dei Experimental Station, CSIC, Zaragoza, Spain
Bio-protocol author page: a2215
, Ernesto Igartua
Ernesto IgartuaAffiliation: Department of Genetics and Plant Production, Aula Dei Experimental Station, CSIC, Zaragoza, Spain
Bio-protocol author page: a2216
 and Frank Ordon
Frank OrdonAffiliation: Institute for Resistance Research and Stress Tolerance, Julius Kuehn-Institute, Quedlinburg, Germany
Bio-protocol author page: a2214
,  date: 5/20/2015, 2640 views, 0 Q&A
DOI: https://doi.org/10.21769/BioProtoc.1473.

[Abstract] Single Nucleotide Polymorphisms (SNPs), which constitute single base-pair variations in the DNA sequence, are the most abundant molecular markers in plant and animal genomes. They are becoming the markers of choice for genotyping in all fields of molecular biology, as they are easily prone to automation ...
1 2 
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