First you get a genomic library of orchid from this atleast you need  a known region(near to ur gene of interest ) of ur genome(example  microsetelight,  snp or RFLP etc)

To this known region u can design a forward and reverse primer.

Amplify it

Design a prob for this known region 

Find where it is in the (Genomic lib )plated colonize 

Took the colony of a known fragments 

(If you can do the  sequencing for the long fragment ) otherwise do the restriction mapping 

Find out end fragment(small one)

Sequences it

Design ur second prob 

Find the fragment in the genomic library 

Likewise repet it till u find the intrested region 

from this u can analysis up words or down words to find the gene of interest

it is called chromosome walking  or chromosome jumping mtd for genome analysis 

Hi Maharajah,

I have around 300 loci that i'm interested in looking at. So this might take a lot of time. But I'll refer more about chromosome walking. Thanks for the suggestion.

E

Hi Eranga,

I would suggest contacting the folks at the 1KP ("1000 plant transcriptomes") for access to Orchidaceae transcriptomes (they have seven right now, it looks like). After you've identified and aligned the regions of interest, you should be good to go!

I've attached a paper that may be of interest for your project.

https://sites.google.com/a/ualberta.ca/onekp/ 

best wishes, Keir

Its ok Eranga If u hv 454 illumina sequencer facilities go with that...

Hi Keir,

Thanks a lot for the suggestion and the links. I'll check them out!

E