Questions tagged [dna-sequencing]

Technique(s) by which the sequence of DNA is obtained. The principles are similar for RNA-sequencing.

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100 views

Why is GenBank growth slowing down?

https://www.ncbi.nlm.nih.gov/genbank/statistics/ shows the growth of the GenBank database is slowing since WGS (Whole Genome Shotgun) emerged. Is this happening because sequencing centers are ...
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How to get phased haplotype from vcf file?

Although I'm reading all days and nights, still confused in this area. I have hundreds of BAM files (from whole-genome sequencing of a given human population). I was wondering how I can obtain phased ...
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totaling OTU from 2 or more observations and assign to new observation in OTU table from qiime

I am new to microbiome processing pipeline but I want to ask, How do I combine two observations and assign it to a new single observation in the OTU table resulted from QIIME. Here is the situation. ...
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How to determine which is the nucleotide sequence of a gene?

I have came across a few genes that show different nucleotide sequence in different databases. I then found out that the sequences are actually reverse complement of each other. How do i determine ...
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Pyura DNA extraction

I'm am struggling with genomic DNA extraction from different samples of Pyura chilensis; the DNA is degraded as can be seen on the gel. We've always used GeneJET Genomic DNA purification Kit (by ...
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If we sequenced the genome of every species, would all phylogenies agree?

The Tree of Life is still up for debate. Most of this debate seems to be due to a lack of genomic information, but that deficiency is decreasing rapidly with advances in technology and sequencing ...
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DNA Sequencing - Sanger termination method: What effects will more ddNTPs in solution have on the resolution on agarose gel via electrophoresis?

Say instead of a small amount of ddNTPs, you add a significant percent of ddNTPs to dNTPs. I was thinking this would reduce the resolution for larger fragments due to more ddNTPs terminating ...
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In a Sanger sequencing, what would be happen if different DNA sequence binds to same primer? And how this situation avoided if problematic?

Sanger's PCR chain-termination sequencing, by its theory, is a simple method to sequence DNA from organisms. But what will happen if there are 2 allelic DNA with same primer binding site, but ...
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2answers
415 views

What direction is a sequence in databases written?

In many databases, the DNA sequences for proteins are given as a string of a,t,g,c without specifying whether the starting is from 5' or from 3'. Also it is not specified if it is the coding or non ...
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Question regarding stop codon

I just wanted to ask that in a given RNA, If there is a start codon and no stop codon, is it still considered mRNA, I mean can it be translated to a protein ?
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Why can't we directly use the DNA instead of the RNA in a DNA microarray analysis? [closed]

When proceeding to a DNA microarray analysis, I saw that the molecule that's taken from the cell is the RNA, which is afterwards converted to DNA by a reverse polymerase. But why not taking directly ...
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Cutting of DNA strand [closed]

Would EcoRI cut the following sequence?? 😕 5'CTCGAGTTCGAG3' 3'GAGCTCAAGCTC5' What is the logic of cutting? Please explain how restriction enzymes work.
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Is there any way to tell if a gene is functional for an individual with only a reference sequence and the individual's sequence of the gene?

I have gene sequence of a ton of individuals within a subspecies from cdna and the ncbi IDed sequence as well. Is there any way to tell using just the sequence that the gene for an individual is non-...
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Dealing with Repetitive Bases in DNA?

I have been given a project involving a plasmid that contains long stretches of Adenine (60 or 120 bases each). These PolyA stretches are interrupted by the occasional G or C. I understand that ...
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2answers
160 views

Complete human rDNA sequence

I've been trying to retrieve the complete human rDNA sequence (non-spacers and spacers), which should be about 43-kb in length using Biomart, NCBI, and rnacentral, but I have only been able to find ...
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Is DNase treatment necessary before RNA sequencing?

Is it necessary to treat total RNA with DNase before sequencing it? In particular, if the library prep relies on a poly-A enrichment, is it necessary to remove DNA, knowing that genomic DNA does not ...
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Is my Y-chromosome the same at 78 as it was at 18?

How accurate is a Y-dna test at age 78?
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Question about phylogenies and DNA sequencing

I'm reading this web page which talks about phylogenies and DNA sequencing: https://bioinf.comav.upv.es/courses/biotech3/theory/phylogeny.html A certain sentence says, "If we consider dolphins and ...
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How do you sequence a tree genome? [closed]

I'm writing a grant proposal, and part of the research will involve sequencing and comparing DNA from trees in America and Japan. How does one analyze the genome of a tree and compare it to a ...
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DNA Longevity in Keratin, Ivory and other Bioplastics

For how long is DNA viable for extraction and/or sequencing in bioplastics created by the human body? I checked out a few links online, and this article gives a record of about 7000 years for frozen ...
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Can PCR duplicates have complementary sequences?

It is common when analyzing paired-end whole-genome shotgun sequencing data to check for and eliminate PCR duplicates. The reasoning is that the probability of sampling a fragment of the genome of the ...
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1answer
1k views

How is DNA testing used to differentiate between different species?

How is DNA testing used to determine that specimens belong to different species, rather than being subspecies, or members of the same species? Usually the definition of species is about the ability to ...
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1answer
358 views

Limitations of Gibson Assembly method

Why is the Gibson assembly method limited to ~8 fragments and what can be done to raise this limit? Why is the method called "seamless" and what are the advantages compared to annealing ...
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Repeating sequences of genomes

How can we estimate what portion of a genome is repeating sequences without sequencing it? Thank you
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PCR products with no band in gel

I am a PhD student and stuck with the no band in the gel results. I am using the following 1 microliter Primer R (20mM) 1 microliter Primer f (20mM) 1 microliter dNTP (10mM) 1 microliter DNA ...
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DNA as a digital storage medium; sequences algorithmically avoided for safety reasons (or should be)?

Safety for the environment is probably implicit here as well, but the focus should be on the people who may come in contact with large amounts of synthetic DNA used to encode information as a data ...
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Replication cohorts in microbial GWAS

Replication in an independent cohort is of course the gold standard in GWAS studies, and many high profile journals will now (quite rightly) not accept finding indicating a phenotype genotype ...
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What does it exactly mean to sequence the genome of a species?

Since all organisms are different from each other - that is, we all have different alleles, some SNP, microsatellites, etc. -, what does it exactly mean, for the genome of a species, to be sequenced? ...
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What are the exact reason causing missing calls in a VCF file from exome sequencing?

My data is a VCF file from exome sequencing variant call. I'm not very familiar with the sequencing process and variant calling process. I noticed that there are some missing genotypes, which is ...
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1answer
225 views

What is the difference between contig- and read-based sequence alignment?

I am trying to understand the difference between read based and contig based alignment. Is contig based alignment refer to de novo assembly and then it is align to a reference genome. I am confused ...
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2answers
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Is it possible to obtain single strands of DNA in solution? [closed]

Being that it is relatively simple to extract DNA from a variety of sources, what further technique can be used to separate the two complementary strands in solution?
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1answer
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How does a polymerized TruSeq adapter affect sequencing reads?

I am trying to build a NGS library on enriched DNA (DNA length = 93 bp). I am using PCR amplification with two overhanging primers, both containing Illumina adapter sequences (Universal Adapter & ...
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1answer
195 views

What are the effects of buffer on genomic DNA shearing by sonication?

Expt: I have isolated genomic DNA from LNCaP cells and would like to crosslink a small molecule (Chem-seq) to the DNA. The small molecule is an alkylator so it will crosslink DNA on its own. The ...
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1answer
87 views

What does it mean to combine in-solution capture of chosen RAD tags to target sequencing reads to desired loci?

The article about "rapture sequencing" found here says: "This protocol also recovers more unique (nonclonal) RAD fragments, which improves both standard RAD and Rapture analysis. Rapture then uses ...
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365 views

Quantifying DNA in a band on gel electrophoresis

I am a first year science undergraduate and I am just asking this as a more theoretical question rather than for carrying out a protocol, so I would appreciate if answers do not involve lots of ...
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2answers
560 views

What are the pros & cons of site-directed mutagenesis? What are the alternatives?

For a lab course we have been assigned to "remove a stop codon, using mutagenesis, that is in the middle of a gene in order for the full gene to be expressed. This will then fluoresce." To do this we ...
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How can DNA profiling be used to determine the number of organisms from a specific species that is within a certain area?

Here is the tedious exam question that asks this: question 8(b). https://drive.google.com/drive/folders/0B23NLD5L6099VWJya3dfLVJpbmc The question states: In 2007, the brown bear was a threatened ...
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157 views

16s rRNA Sequencing From Gut Microbiome (stool)

Do I extract RNA or DNA from gut microbiome (stool samples) if want to do 16s rRNA sequencing?
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How do we know which Eulerian path corresponds to the DNA sequence?

Bioinformatics textbooks explain one method to reconstruct the whole DNA sequence from multiple reads. The method builds a graph and finds a Eulerian path in it. The Eulerian path corresponds to a DNA ...
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2answers
347 views

What does chrUn mean in the output from a Bowtie run on human sequences?

After having performed an alignment using bowtie2 and GRCh38 as a reference sequence, I got inusual matches on chrUn. Here a small part of the SAM file: ...
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1answer
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Why is only one primer used for DNA sequencing instead of two?

I understand that PCR uses two primers that anneal to the two ssDNA's in order to exponentially amplify a DNA and that Sanger sequencing uses only one primer because a sequence can be determined with ...
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Finding protein motifs by running sequence analysis in protein BLAST

I've run the following sequence (bottom of page) in NCBI protein BLAST and its been identified as Zn-dependent alcohol dehydrogenase. Next I want to see what protein motifs are in this protein. Where ...
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2answers
226 views

Actual determination of the DNA sequence in the shotgun approach?

I'm studying bioinformatics and I'm confused by shotgun sequencing. In Sanger sequencing we break up the DNA and use ddNTPs in order to determine the exact position of each neucleotide. How exactly ...
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1answer
59 views

How to find sequencing data of a specific drosophila strain

I'm trying to correct low coverage parts of an assembly of these PacBio reads with Illumina data. According to PacBio they "have sequenced adult males from a subline of the ISO1 (y; cn, bw, sp) strain ...
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An efficient and free method of assembling a large number of Sanger sequencing reads?

I have a rather large number of Sanger Sequencing reads.  Is there a automated method in either Ugene or a similiar freeware that allows you to quickly assemble forward and reverse reads. Ideally ...
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Where can I get my exomes sequenced?

I am a private individual and I want my exomes sequenced, I don't know of any place that will keep my data confidential as I read reports of companies sharing this data with insurance companies. I ...
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Are SNPs and alleles the same thing?

It seems to be quite difficult to find an answer to this. Are SNPs the same thing as alleles?
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Can DNA sequencing of food provide quantitatively the fraction of protein from chicken vs soy?

There is some news about a report from Canada's CBC about fast-food chicken items containing as much as 50% soy protein, based on DNA testing. I won't do the report the service of linking to it ...
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VCF files - examples for haploid organisms

I am trying to self-learn some aspects of bioinformatics (coming from a stats background), and i am trying to find / download some VCF files for haploid organisms as an input to some statistical ...
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How to design a ChIP-exo experiment with controls?

Does anyone have any ideas on how to run a control/input sample for a ChIP-exo experiment? Is it even necessary to run a control (similar to input DNA for ChIP-seq)? I'm looking for any references ...