Hi Silvio,

Design a primer from the backbone, and a primer from your gene of interest for PCR. This can verify (1) whether your gene is inserted, (2) the orientation of your inserted gene, and (3) the PCR product can be used for sequencing.

To check the transgene copy numbers using Southern, you should design primer(s) based on your transgene sequence.

Thanks for your answer. My problem is that I don´t found the vector sequence. Probably pMON546 is a derivative of M13 vector but I am not sure, and the vector sequence is missing in NCBI. Any information about this is very useful for me.

Thanks again.

Silvio

Hi Silvio,

You might try to make a phone call (custom service) to the company (ATCC) below to ask--to see whether they can provide you the sequence of pMON546. They are selling this plasmid. On the website, the custom service phone #: 1.800.638.6597

http://www.atcc.org/products/all/53213.aspx#documentation

There also one reference on the page: Shah DM, et al. Glyphosate-resistant plants. US Patent 4,940,835 dated Jul 10 1990. It may be useful for you to take a look. [ http://www.google.com/patents/US4940835 ]

Attached is an article. Please see a simple diagram/plasmid map and explanation of pMON546 on the pages 91 and 92. I hope it will offer some help.

Thank you so much. Your information is very useful for me.