Questions tagged [dna-sequencing]

Technique(s) by which the sequence of DNA is obtained. The principles are similar for [tag:rna-sequencing].

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Basic text/notes on DNA structure (for non-biologists)

I am a mathematics grad student researching knot theory, and I've recently discovered that there is a connection between knot theory and DNA structure (if I understand correctly, when DNA strands ...
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Why only heterogeneous SNVs for validation using genotyping arrays?

I am trying to validate the variants I found using whole genome sequencing . The standard practice, I have seen in the two publications below were to check for the number of heterozygous SNPs called ...
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How to calculate extent of Sequence similarity [closed]

I had a problem that I was wondering if it could be solved by one of the techniques/algorithms used in bioinformatics to give the extent of similarity. I have a Problem Statement: we have a sensor (...
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4answers
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What species have had their genomes sequenced/are being sequenced?

The human genome project released it's first complete genome nearly ten years ago. Since then many species have also been sequenced. I am trying to find a list of completed (and possibly ongoing/...
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Is it possible to clone a Neanderthal?

As Neanderthal DNA has been sequenced, is it possible to create a living Neanderthal?
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276 views

DNA preparation for sequencing

In shotgun sequencing method or some related method that DNA is break up into random fragment. The fragment that have about 3kb in size is inserted into plasmid by enzyme ligase and then plasmid ...
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Why can't we observe haplotypes directly?

I am completely a noob to biology and DNAs, but recently I am looking into EM-algorithm used in the case of re-constructing haplotypes from genotype data. I am just wondering what exactly in the ...
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1answer
235 views

Any tool to align whole genome sequence data to another genome and give exon regions a higher mark?

I want to align WGS data from several mammals to one reference such as the human genome sequence. Since in most cases exons should be conserved and spliced in the same way and introns should vary, I ...
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Can Pfx polymerase add only one 3' A overhang?

I am trying to clone a PCR product that was amplified using Pfx polymerase into pGemT vector. I had to A-tail the PCR product using Taq polymerase since Pfx only generates blunt end products. My ...
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90 views

Are there beneficial genetic mutations identified by consumer DNA genotyping?

I'm looking at services like 23andme, and see that they identify a wide variety of genetic-based risks, like predisposition to diseases, hair loss, cancer, etc. Are there a more "positive" DNA ...
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Reverse complement of reconstruction model for assembling reads

One way to assemble fragments produced by DNA sequencing (often called reads) is to seek for the shortest common superstring that contains all the reads of a given set of reads. One model for this ...
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1answer
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What is the purpose of Y-shaped adapters in Illumina sequencing?

Y adapters different sequences to be annealed to the 5' and 3' ends of each molecule in a library. The arms of the Y are unique, and the middle part, connected to the DNA fragment, is complementary. ...
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Why is Sanger sequencing inferior for detecting SNPs in cancer cells?

I am familiar with Sanger sequencing, but at the level of an undergraduate. A lecturer of mine tried to describe Sanger sequencing as losing the sequence information in noise when used to detect ...
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What is solid-phase bridge amplification?

During Illumina sequencing there is a step called bridge amplification by which DNA is amplified by isothermal enzymes. What is this stage, and how does it work?
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What's the difference between shotgun sequencing and clone based sequencing?

In a lecture during my undergraduate degree we were introduced to the race to complete the human genome. Celera were competing with Sanger and collaborators to sequence the human genome. Celera ...
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BLAST DNA Sequences Reversed

I have been trying to learn some basic DNA sequencing techniques and have been using BLAST as a reference. I thought that I was starting to get it, but then I cam across this: It looks like it's ...
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3answers
310 views

Results of a complete DNA sequencing - are they 100% reusable?

Is that correct that a complete DNA sequencing (the whole genome) need only to be done once (per person)? After that is done, it the complete genome can be stored and once the new genes (and their ...
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Separating DNA Fragments by Gel Electrophoresis. Are all the strands for one size the same?

My apologies if my question is too basic, and please point me to a more appropriate forum. I am reading the textbook "Essential Cell Biology" by Alberts et al, and am consulting other sources as well....
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Tool for nucleotide alignment with all nucleotide codes (e.g. R, Y, W, S, etc.)?

I have a vector sequence and would like to find the following nucleotide sequence in it. AASYWSRA This query sequence uses several degenerate symbols, defined as:...
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How are geneticists able to isolate gene functions?

As an example, this Scientific American article describes a known area in the dog genome that metabolizes carbohydrates. How is it that researchers are able to determine specific functions such as ...
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Sequencing rtPCR product

So I have a validated set of primers for rtPCR from Biorad that contains SYBR green. If I do rtPCR, can I use the rtPCR product after purifying it with a Qiagen PCR purification kit? Also, I'm under ...
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1answer
5k views

Number of reading frames in nucleotide sequence

In class of molecular biology was posed this question: "How many reading frames start in sequence of nucleotides 5' ... ATATGAATGATGACAT... 3' ?"
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What are the limitations of commercial-grade DNA genotyping compared to full sequencing?

I've heard about services like 23andme, which offer genetic testing to the general public. As a person who knows very little about genetics, I'm interested in the subject and would like to know what ...
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3answers
2k views

Finding proteins in DNA sequence

I have to do a task for a university task and I need to understand some things before figuring out how to do it. The task is the following: Find matches of known proteins (DNA-PolyI,II,III) to the ...
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What is the difference between SOLiD, 454, and Illumina next-gen sequencing?

I've started teaching myself about next-generation sequencing in preparation for a new job, and I'm wondering what the main differences are between the 454, SOLiD, and Illumina/Solexa machines, in ...
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Comparative cost of RNA-seq vs sequencing full length cDNAs

I am in the process of assembling and annotating the genome of a non-model organism, using almost exclusively short read (paired-end Illumina) data. Throughput is one obvious benefit of these data (...
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1answer
1k views

Digital Genomic Footprinting for ENCODE

I'm reading over the ENCODE Nature papers, and one of the papers referred to is "Global mapping of protein-DNA interactions in vivo by digital" by Hesselberth et al[1]. Genomic footprinting is a ...
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Sequence of ribosomal RNA

Is it possible to sequence rRNA directly, that is, using the ribosome rather than the DNA from the nucleus? For example, this paper, Complete nucleotide sequence of a 16s rRNA gene from E. coli, ...
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341 views

Is there a detectable amount of bacterial DNA in the blood of infected persons?

With which bacterial infection in humans has it been shown that bacterial DNA can be found in the blood? If any is found it is likely not to be very much, and even difficult to distinguish from ...
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225 views

Sequencing from PCR

As far as I understand it, PCR can be used to make many copies of one gene. My question is, is it possible to sequence DNA after PCR and is it easier than sequencing it via other methods. If it is ...
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1answer
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Is it possible to sequence only the 16S rRNA component? If so, how?

There have recently been several papers on using 16S rRNA as a way of identifying species (here, and here). I'm wondering if it's possible to sequence either just that subunit of the ribosome or just ...
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Why did high A+T content create problems for the Plasmodium falciparum genome project?

The main paper for the Plasmodium palciparum genome project (Gardner et al., 2002) repeatedly mentioned that the unusually high A+T content (~80%) of the genome caused problems. For example they imply ...
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1answer
790 views

ChIP-seq vs ChIP-exo

I'm currently investigating ChIP-seq vs. ChIP-exo for finding binding sites. As far as I can tell, ChIP-exo seems to be better in every way than ChIP-seq... but then again, I'm not strong in this ...
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Can I compare Shannon indices of metagenome gene data?

I'm comparing 12 Metagenomes. I'm using HMM counts of a number of proteins known to exists as groups within certain operons. I have grouped the HMM counts for each type of operon and calculated the ...
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Is there any difference in terms of personal healthcare between complete DNA sequencing and SNPs genotyping?

There are several companies (most notably 23andMe) providing SNP genotyping for individuals claiming that there is no sense for individual in obtaining full genome sequence since only small part of it ...
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What's the use of DNA sequencing results?

Suppose I sequence DNA of some organism (a human perhaphs) and now I have the entire DNA "string" - the sequence of nucleotides. What's the use of that? It's just a "string" where nucleotides encode ...
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4answers
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What exactly are computers used for in DNA sequencing?

I've thoroughly read the Wikipedia article on DNA sequencing and can't get one thing. There's some hardcore chemistry involved in the process that somehow splits the DNA and then isolates its parts. ...
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Why are there N's after Sanger sequencing?

After sending a DNA sample for sequencing, the resulting sequence had N's in the beginning and end of the sequence. I know the N's mean that the computer can't tell what the base pair is, but why is ...
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1answer
153 views

Mechanics of going from DNA sequence to metabolic network

My apologies for possibly a naive question. I'm a statistics type and have been asked to assist with analysis of metabolic pathways (and networks). However, I would like to have at least a layman's ...
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protocol for pulldown of DNA breakpoints?

Is there any method to do pulldown enrichment of DNA breakpoints from a cell? I have found this paper reporting a method to enrich for the DNA single-strand breakpoints from meiotic recombination ...
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1answer
2k views

How to clone and sequence a gene transcript of unknown sequence?

How might I go about amplifying a gene transcript (mRNA) from animal tissue of which little is known about the genome? In some applications, I have used reverse transcriptase PCR to amplify all mRNA ...
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How to create a collection of anonymous sequences for teaching and testing? [closed]

I am looking for a large collection (>1000) of sequence files (eg. FASTA) from any real organism or a tool to create such a collection. The sequence files would be used for teaching and for testing ...
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Why do restriction enzymes tend to have an even number of bases in their recognition site?

When reading my textbook I noticed that in all examples but one from eight the recognition site was an even number of bases. I wondered if this was just a co-incidence, so I took the data from this ...
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698 views

Questions to ask to a panel of people that will be sequenced

Some genes have been shown to be associated with left-handedness. Working with some clinicians, I've recommend them to ask their patients (whose genome will be sequenced) if they are "right or left-...
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1answer
986 views

Sequencing the genomes of polyploid organisms

I've done some transcriptomics work in the past with a polyploid organism, and this presented some unique challenges in the data processing and analysis. Since then, I have been brainstorming about ...
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What are the limitations to current nucleotide sequencing technologies?

Using the Illumina platform, it is cheap and (relatively) easy to sequence huge amounts of DNA or RNA. There are various other platforms out there (Roche/454, SOLiD, PacBio, Ion Torrent) each with ...
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1answer
75 views

How are whole Haplotypes for Sequencing isolated?

Is it possible to reliably isolate and amplify DNA from individual sperm and/or eggs (from a fish)? I'd imagine that the small amount of DNA would make the PCR a bit wacky. We've considered ...
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2answers
643 views

Looking for a cancer drug target database to guide sequencing of patient tumor DNA

I have a question I would like to pose to the community. I have recently received access to a bench-top ion torrent DNA sequencer. Our idea is to use this machine to sequence the DNA from patient’s ...
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476 views

Why was it so hard to decode the corn genome?

My teachers growing up told me it was impossible to decode the maize genome. But yet its been done. Why was decoding the genome so significant, and what made it so difficult?
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Alternate genetic codes in newly sequenced organisms

Variations of the standard genetic code are pretty rare, but as the cost of high-throughput genome sequencing continues to drop, there is a greater possibility of discovering additional exceptions. ...