Blue:RNA-Seq Experiments:08122013

From ZhangLabWiki
Jump to navigation Jump to search

Purpose[edit]

  • Sort single huES8 (Alb iPA) stained positive for SSEA4 per well for totoRNAseq analysis

Prepare RT buffer:[edit]

' Stock Final (5x) Vol for 1 ml
Tris-HCl pH8.3 1M 250mM 250ul
KCl 1M 375mM 375ul
MgCl2 1M 30mM 30ul
dH2O 315ul


Prepare RT Sort Buffer:[edit]

Component Volume
5xRT Buffer 250ul
10% Triton-X100 5ul
40U/ul Rnase Inhibitor 12.5ul
ERCC spike-in RNA (1:10^5) 5ul
Water 227.5ul
total 500ul
Note: using 2ul per well


Prepare Quartz-Seq Sort Buffer:[edit]

Component Stock Final (1x) Volume
dH2O 191ul
Triton-X100 10% 0.1% 2ul
Rnase Inhibitor 40U/ul 1U/ul 5ul
ERCC spike-in RNA 1:10^5 2ul
total 200ul


Note: using 0.4ul per well and generating bubbles just prior to sort

Plate Layout:[edit]

' 1 2 3 4 5 6 7 8 9 10 11 12
A NC(1) UHRR(9) NC(17) NC(25) UHRR(33) NC(41) NC(49) UHRR(57) NC(65) NC(73) UHRR(81) NC(89)
B Sc(2) Sc(10) Sc(18) Sc(26) Sc(34) Sc(42) Sc(50) Sc(58) Sc(66) Sc(74) Sc(82) Sc(90)
C Sc(3) Sc(11) Sc(19) Sc(27) Sc(35) Sc(43) Sc(51) Sc(59) Sc(67) Sc(75) Sc(83) Sc(91)
D Sc(4) Sc(12) Sc(20) Sc(28) Sc(36) Sc(44) Sc(52) Sc(60) Sc(68) Sc(76) Sc(84) Sc(92)
E Sc(5) Sc(13) Sc(21) Sc(29) Sc(37) Sc(45) Sc(53) Sc(61) Sc(69) Sc(77) Sc(85) Sc(93)
F Sc(6) Sc(14) Sc(22) Sc(30) Sc(38) Sc(46) Sc(54) Sc(62) Sc(70) Sc(78) Sc(86) Sc(94)
G Sc(7) Sc(15) Sc(23) Sc(31) Sc(39) Sc(47) Sc(55) Sc(63) Sc(71) Sc(79) Sc(87) Sc(95)
H Sc(8) Sc(16) 10cell(24) Sc(32) Sc(40) 10cell(48) Sc(56) Sc(64) 10cell(72) Sc(80) Sc(88) 10cell(96)


Sc=Single cell; (#)=Indx