It would be difficult to clone a 9.7 kb fragment in TA cloning plasmid in one step. I suggest you either to divide it into 2-3 fragments, clone them then reclone them to make into one piece. Or try other strategies, like infusion or USER or Gipson cloning strategy, which can assemble multiple fragments with one go.

Hi, there are some kits specific for long amplicon, for example:

StrataClone PCR cloning kit has been used to clone PCR products up to

9.2 kb in length http://www.genomics.agilent.com/en/PCR-Cloning-Kits/StrataClone-PCR-Cloning-Kits/?cid=AG-PT-161&tabId=AG-PR-1190.

You can try this or search on the web

Simple TA cloning kit may not handle larger fragments (>3kb);

For larger fragments (3-10kb); Topoisomerase  based TA kit (TOPO® XL PCR Cloning Kit) is available with LifeTech. It will help you

http://www.lifetechnologies.com/in/en/home/references/protocols/nucleic-acid-amplification-and-expression-profiling/pcr-protocol/topo-xl-pcr-cloning-kit.html

thanx to all for suggestions

Dear Chandrajeet,

TOPO XL-2 can work with up to 20 kb fragments. I doubt there is an enzyme which will work for you for such long fragments. You may try Plat Superfine (Life Technologies) or Q5 HS HiFi (NEB). Do not try KOD systems as the enzyme does not produce blunt ends and gives poor results during cloning (poor proofreading). The third option is a Takara system and it works supposedly up to 30 kb fragment length. The system worked wonders for me when everything else failed.

Best regards,

Wes