Cloning inserts: forward and reverse orientation - (Sep/18/2014 )

good day

If a insert is in the reverse orientation does that mean the anti sense strand is now the sense strand or does it just mean the sequence is now read from back to front, when compared to its forward orientation?

EXAMPLE:  FORWARD:      5' ATCGGG 3'

                                              3' TAGCCC 5'

IS REVERSE THIS WAY : 5' GGGCTA 3'

                                           3' CCCGAT 5'

OR THIS WAY 5' CCCGAT 3' 

                         3' GGGCTA 5'

THANKS

The bottom one. Note where the 5' and 3' labels are. You can simply lift the top sequence up, flip it right to left and top to bottom. The cell and chemistry don't know you have done that.

phage434 on Thu Sep 18 14:21:15 2014 said:

The bottom one. Note where the 5' and 3' labels are. You can simply lift the top sequence up, flip it right to left and top to bottom. The cell and chemistry don't know you have done that.

but then wouldn't reverse orientation change everything, example, revering to my initial questions example: if ATCGGG was a enzyme recognition site, now in reverse it is CCCGAT. The enzyme wouldn't recognize that? 

That site is still present in the bottom picture. Chemistry and biology look at the location of the 5', not where things are printed on a page. If you look at the bottom picture, and examine the bottom strand, you'll see the 5' on the bottom right. Now read (in the 5' to 3' direction) towards the left, and you'll see the sequence (written 5' to 3') ATCGGG, which is the same sequence (again, written 5' to 3') as the top strand on the original picture.

phage434 on Fri Sep 19 12:55:10 2014 said:

That site is still present in the bottom picture. Chemistry and biology look at the location of the 5', not where things are printed on a page. If you look at the bottom picture, and examine the bottom strand, you'll see the 5' on the bottom right. Now read (in the 5' to 3' direction) towards the left, and you'll see the sequence (written 5' to 3') ATCGGG, which is the same sequence (again, written 5' to 3') as the top strand on the original picture.

oooooh I see now, thanks