Question: Forward And Reverse Strand Conventions
18
gravatar for Andrea_Bio
4.8 years ago by
Andrea_Bio2.2k
Andrea_Bio2.2k wrote:

Hi

I apologise for the really basic question. I've probably said this a million times now but I'm returning to this field after a long time and I keep confusing myself by mis-remembering or half-remembering things from the past and it's not helping me. It would be easier coming to the field from fresh i think. Anyhow...

As i remember it, sequence databases always store the forward strand of a DNA chromsome in the 5' to 3' direction.

A gene is read in the 3' to 5' direction and so its complementary strand in the 5' to 3' direction is the same as the mRNA transcript. So i thought if a gene was on the reverse strand of a DNA molecule then the forward strand in the 5' to 3' direction gives the sequence on the corresponding mRNA (ignoring introns for simplicity).

However I'm just looking at a gene now in ensembl and this gene is described as being on the forward strand. If you look at the forward strand it contains the exact same sequence as the gene's mRNA. So to me then, if the mRNA runs on the forward strand in the 5' to 3' direction the actual gene is on the reverse strand is it not?

Is this a convention issue that I have mis-remembered? Is a gene classed as being on the forward strand if its mRNA sequence is 'on' the forward strand.

thanks for your help

sequence strand • 45k views
ADD COMMENTlink modified 4.6 years ago by Neilfws45k • written 4.8 years ago by Andrea_Bio2.2k
30
gravatar for Bio_X2Y
4.8 years ago by
Bio_X2Y3.1k
Ireland
Bio_X2Y3.1k wrote:

Sounds like you're trying to knit a few hazy concepts together too quickly - it might help to start by completely ignoring the RNA polymerase (the machinery that does the transcription in the "opposite" direction), and building up your understanding from scratch. Here is a stab at an explanation, hopefully it won't make things worse!

Start with the basics:

  • DNA is double-stranded. By convention, for a reference chromosome, one whole strand is designated the "forward strand" and the other the "reverse strand". This designation is arbitrary. Sometimes the terms "plus strand" and "minus strand" are used instead.

  • Visually (I'm not talking about the transcription machinery yet), you would typically read the sequence of a strand in the 5-3 direction. For the forward strand, this means reading left-to-right, and for the reverse strand it means right-to-left.

  • A gene can live on a DNA strand in one of two orientations. The gene is said to have a coding strand (also known as its sense strand), and a template strand (also known as its antisense strand). For 50% of genes, its coding strand will correspond to the chromosome's forward strand, and for the other 50% it will correspond to the reverse strand.

  • The mRNA (and protein) sequence of a gene corresponds to the DNA sequence as read (again, visually) from the gene's coding strand. So the mRNA sequence always corresponds to the 5-3 coding sequence of a gene.

  • Now, the RNA polymerase machinery moves along the DNA in the 5-3 orientation of the coding strand (e.g. left-to-right for a forward strand gene). It reads the bases from the template strand (so it is reading in the 3-5 direction from the point-of-view of the template strand), and builds the mRNA as it goes. This means that the mRNA matches the coding sequence of the gene, not the template sequence. (This diagram from Wikipedia illustrates).

  • Annotations such as Ensembl and UCSC are concerned with the coding sequences of genes, so when they say a gene is on the forward strand, it means the gene's coding sequence is on the forward strand. To follow through again, that means that during transcription of this forward-strand gene, the gene's template sequence is read from the reverse strand, producing an mRNA that matches the sequence on the forward strand.

ADD COMMENTlink modified 14 months ago • written 4.8 years ago by Bio_X2Y3.1k
1

Also trying to get this straight in my head. @Bio_X2Y:

you mention

"the RNA polymerase machinery moves along the DNA in the 5-3 orientation of the coding strand ... and builds the cDNA as it goes."

Not trying to be obtuse here, but doesn't the RNAP make RNA which would be complementary to the template which is identical to (except U's instead of T's) the coding strand?

"This means that the cDNA matches the coding sequence of the gene, not the template sequence."

Wouldn't the cDNA then, which is complementary to the mRNA (and coding sequence), be identical to the template?

ADD REPLYlink written 3.5 years ago by Onefishtwofish10

I thought that was exactly what i said in my question? I quote: Is a gene classed as being on the forward strand if its mRNA sequence is 'on' the forward strand.

I've just always called the coding strand the mRNA sequence. Like you say the template strand, which to me is the actual gene, is on the reverse strand. I should have been clearer wih my wording

ADD REPLYlink written 4.8 years ago by Andrea_Bio2.2k

forgot to say thank-you!

ADD REPLYlink written 4.8 years ago by Andrea_Bio2.2k

Fair enough, I guess the answer is "yes" then :) I partially used the answer to clarify things in my own head too. I can understand why you consider the template strand to be "actual" gene, but I would always have conceptualised it as the other way around (feels easier to me anyway). Things must get interesting in your view when you consider anti-sense transcription :)

ADD REPLYlink written 4.8 years ago by Bio_X2Y3.1k

Great answer, thanks for putting the time into crafting a detailed explanation!

ADD REPLYlink written 4.8 years ago by Istvan Albert ♦♦ 56k

@Bio_X2Y : Your first mention that the designation of forward and reverse strands is arbitrary. Are you sure about this ? I imagined that the forward strand was the one with the 5' end closest to centromere, no ?

ADD REPLYlink written 4.7 years ago by toni2.0k

@tony, I think you're correct - I ended up posting a follow up question here: http://biostar.stackexchange.com/questions/3916/conventions-for-designating-forward-and-reverse-strands

ADD REPLYlink written 4.6 years ago by Bio_X2Y3.1k

Also trying to get this straight in my head. @Bio_X2Y:

you mention "the RNA polymerase machinery moves along the DNA in the 5-3 orientation of the coding strand ... and builds the cDNA as it goes."

Not trying to be obtuse here, but doesn't the RNAP make RNA which would be complementary to the template which is identical to (except U's instead of T's) the coding strand?

"This means that the cDNA matches the coding sequence of the gene, not the template sequence."

Wouldn't the cDNA, which is complementary to the mRNA (and coding sequence), be identical to the template?

ADD REPLYlink written 3.5 years ago by Onefishtwofish10

Maybe I am misunderstanding the answer...

Quote: "the RNA polymerase machinery moves along the DNA in the 5-3 orientation of the coding strand ... and builds the cDNA as it goes." Not trying to be obtuse here, but doesn't the RNAP make RNA which would be complementary to the template? Quote: "This means that the cDNA matches the coding sequence of the gene, not the template sequence." I would have thought that the cDNA, which is complementary to the mRNA (and coding sequence), is identical to the template, and not the coding sequence. Am I thinking about this wrong?

ADD REPLYlink written 3.5 years ago by Onefishtwofish10
4
gravatar for Neilfws
4.8 years ago by
Neilfws45k
Sydney, Australia
Neilfws45k wrote:

Short answer - yes. A "gene" is on the forward strand if its mRNA is on the forward strand.

I placed quotation marks around "gene" because, perhaps surprisingly, it is not a very useful word. In the context of this question we're using "sequence of a gene" to mean the same as "sequence of the mRNA transcribed from the gene." So we're referring to the same strand because we're talking about the same object.

What is a gene though? Most "genes" give rise to multiple transcripts. So we might say a gene is a region of DNA that serves as a template for transcription. A gene is not really a single object with start and end. It will have a "minimum start" (the 5'-most base from which transcription occurs) and a "maximum end" (the 3'-most base at which transcription terminates). So perhaps it's best to forget about genes and think about transcripts and their properties.

I agree that the "start < end" convention takes some getting used to but it makes a lot of sense when it comes to performing range calculations with sequences. If you just name the strands "+" or "-" and set start < end, you can forget about all the other terminology (forward/reverse, 5'/3' and so on), it all "just works".

ADD COMMENTlink written 4.8 years ago by Neilfws45k
0
gravatar for Pierre Lindenbaum
4.8 years ago by
France
Pierre Lindenbaum74k wrote:

'...A gene is read in the 3' to 5' direction and so its REVERSE complementary strand in the 5' to 3' direction is the same as the mRNA transcript...'

ADD COMMENTlink written 4.8 years ago by Pierre Lindenbaum74k

THE cDNA sequence always corresponds to the 5-3 coding sequence of a gene. You dont need to reverse it.

ADD REPLYlink written 4.8 years ago by Andrea_Bio2.2k

I meant the genomic sequence of the gene. See the SQL table at the UCSC, all the chromStart positions are lower than chromEnd positions whatever is the orientation.

ADD REPLYlink written 4.8 years ago by Pierre Lindenbaum74k

I don't think i follow what your point is. Bio_x2y has confirmed for me what I thought about template and coding strands.

I think what you are talking about is another conventions issue whereby the start of a gene is always less than end of a gene even if you are on the reverse strand. On the reverse strand the start of the coding strand is higher than the end if you are using the 5' end of the forward strand as base 1. But by convention you give the gene coordinates so start is always less than the end (and flip the start and end coordinates in your transformations)

ADD REPLYlink written 4.8 years ago by Andrea_Bio2.2k

Personally i've always thought it looks a bit odd when you see a gene on the reverse strand and the start position of its second exon is lower than the end positon of its previous exon. I gues you just get used to reading from right to left for reverse strand stuff

ADD REPLYlink written 4.8 years ago by Andrea_Bio2.2k
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