1 Take cells out of -80C and thaw on ice for 5 min. 2  Add 1 ul (~500 ng) plasmid DNA to 50 ul cells, mix gently with pipette tip. 3  Leave on ice for 30 min. 4  Put in 42C water bath for 45 sec.
Melisandre is a red priestess of R'hllor and a shadowbinder, hailing from the city of Asshai in the Further East of Essos. She has joined the entourage of Stannis Baratheon, Lord of Dragonstone, believing him to be Azor Ahai reborn, a hero destined to defeat the Great Other.
Jovelina Farias Belfort, most famously known as Jovelina PÃ©rola Negra (Jovelina Black Pearl) (Rio de Janeiro, July 21, 1944 - November 2, 1998) was an Afro-Brazilian singer. She is considered to be one of the great ladies of samba and pagode with her strong voice, rich in tone and strength.