3'-end labeling of tRNA

[January Weiner]

Hello,

	I am labeling  the 3'-end of tRNA using [32-P]pCp. Or, rather, I am
	planning to do it. 

	Some questions:

	1) do you have a nice protocol for making radiolabelled pCp from
	3' cytidine phosphate? I have a following one, don't know whether
	it is OK:

	speed-vac dry 100uCi gamma-ATP
	+2 ul pCp-mix
		pCp-mix: 3ul 1M Tris ph 8.0
		2.5 ul 0.1 M MgCl2
		1 ul 0.2M DTT
		2 ul 20mM spermidin
	+1 ul 1mM Cp (cytidine 3'-phosphate)
	5U PNK
	---
	30' @ 37 deg, 2'@ 100 deg
	speed-vac dry

	Would you use just standard PNK buffer? (end conc. 50mM Tris 7.5, 10
	mM MgCl2, 5mM DTT, 0.1mM spermidine)

	2) where do you buy 3' cytidine phosphate? 

	Regards,
	j.

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