There are many reasons to clone the DNA fragment in plasmid, few are

1. The bacterial DNA replication have proof reading activity so less chance of error

2. Ease of generating DNA fragment array/library

3. Easy to multiply when ever needed

4. If the DNA fragment is a CDS can be further studied using expression vector

Many researchers in many fields do sequence PCR products directly.  The product is often purified by gel electrophoresis or chromatography before sequencing.   The result is a "cosnensus" of the PCR products, so if you are working with a diploid organism one or two sites may show a double peak (A from mom's allele and C from dad's allele for example).  If you are working with a virus population such as HIV-1 in blood any ratio could be found, not just 50:50 as expected in diploid cells.  Mitochondria genes should be pure, genes from a bacterial isolate/clone should be pure.

You can sequence PCR products directly, just as long as your sequencing primer lies within the region of the amplified fragment.

1. PCR products can be directly sequenced, or it can be cloned into a sequencing vector for sequencing using the specific primers on the vector.

2. If your PCR products contain several species with very little difference (for example in size, 199 bp, 200 bp, 202 bp .etc, due to induced base mutations), which almost cannot separated on the gel, then the gel-purified product from this 'single' band should be cloned into plasmid before sequencing to figure out each species' sequence. One example is creating a null allele through the using of designer nucleases (ZFN, TALEN,...) on plant protoplasts. The disrupted allele (null) can has deletion or addition of only several bases in it. PCR from the DNA isolated from this pool of cells using the allele's primers can contain several mutants.

You can sequence PCR products directly if you have single band or you can purify desire band and can sequence...

Maybe it's just because it's cheaper where we submit our samples for sequencing. Go figure.

Hello,

You can sequence cleaned PCR product directly and you can cloned it before, it depends on what you have and what you want to do.

For exemple if you have heterozygosity and you need to separate alleles or when your PCR product is precious or quantitatively low, it's better to clone it.

Good luck 

There are many reasons to clone the DNA fragment in plasmid, few are

1. The bacterial DNA replication have proof reading activity so less chance of error

2. Ease of generating DNA fragment array/library

3. Easy to multiply when ever needed

4. If the DNA fragment is a CDS can be further studied using expression vector

You do not need to, you can sequence the PCR product directly. But make sure that the amplification has been done using a high fidelity polymerase in the first place.

As my colleagues said above, you can sequence the PCR product directly, but usually it is better to clone it, because of different reasons: the first is because you will have a "stock" of your PCR product for future works on it, the second because usually the first 10-50 bp of the Sanger sequence fragments content a lot of noise, so to be sure that you have the complete sequence of your pcr product, if you clone it on a vector you can perform sequencing using specific primers from the vector, so you could be sure that you will have the complete sequence of the PCR product (depending on the size of it of course). Other reasons have been explained: to detect puntual mutations, you can sequence several clones, etc.

Hi John,

I agree with monica as well as the other comments.

I have sent cloned PCR products as well purified PCR products to be sequenced for the same gene and the cloned samples have far less noise than the PCR products that have been purified and not cloned. 

i also agree with monica

One more thing: Sequencing reaction is optimised for  vector primer (universal primer) so the sequencing result alway better. You also can not obtain sequence near primer because signal sequencing alway not good enough (about 60 nucleotide after primer)

I Agree Completely with the answer of  Bryan Thomas Folly:

Many researchers in many fields do sequence PCR products directly. The product is often purified by gel electrophoresis or chromatography before sequencing. The result is a "cosnensus" of the PCR products, so if you are working with a diploid organism one or two sites may show a double peak (A from mom's allele and C from dad's allele for example). If you are working with a virus population such as HIV-1 in blood any ratio could be found, not just 50:50 as expected in diploid cells. Mitochondria genes should be pure, genes from a bacterial isolate/clone should be pure.

BUT an Excellent reason to put a cDNA in a specific plasmid suchas ones containing SP6 T7 sequences  is the possibility huge quantities of you CDNA allowing you to produce nearly ad libitum RNA probes fro in situ hybridization on whole embryos or on tissue sections

Even if you decide to clone, you can save time save money. we pcr directly from colonies and sequence directly from PCR.   thus you can have cloned DNA and sequence at the same time.

If your future plan is to clone your gene of interest into vector then you can first directly do sequencing of your PCR product (i.e. before cloning) and then after cloning you can confirm it again by doing sequencing using  vector as well as your primers. If your plan is just do PCR and confirm the product by sequencing then you can do only sequencing of your PCR product. 

Hello

If is possible to sequencing PCR directly? Yes, it is possiblre BUT usually you will loss information beacuse about the first 50 to 70 bp does not have good reading. If you have interest in each base pair of you PCR product is necessary  cloning this to vector like pGEM-T. I agree with the others colleagues is better sequencing using primers que aneal on vector sequences.

Hi All,

Just had a following up question. I work on a diploid organism with high heterozygosity. I'd like to figure out the two haplotypes of this organism at some genomic regions. That's why I am doing DNA cloning.

As far as I understand, during ligation process, a single PCR fragment will be ligated to a single vector, is it possible that during transformation process multiple vectors with different inserted PCR fragments (let's say, in my case, two vectors with two haplotypes inserted) get into a single bacterial cell, if so, how would it affect downstream sequencing?

Hope my question was clear.

Thanks!

I run into the same question when checking crispr knock out. And I find useful answers here--

1. From Yuan-Yeu Yau, single-cell colonies make sanger results clear

2. From Monica Martinez and Julio Levano Garcia, sanger always loses first 10-70bp or 50-70bp?