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b******r
发帖数: 111 | 2 I am struggling for testing primers to genotype transgenic mice. I use test
1fg of vector as template mixed with 1ng of genomic DNA,
1. PCR always works when the template contains 1fg of vector but no genomic
DNA.
2.PCR always doesn't work when the template contains both 1fg of vector and
1ng of genomic DNA.
PCR conditions: 25 uL volume, pfu enzyme, 95 for 25",65 for 25",72 for 2'.
product size 1kb
Any solutions to fix this problem? Any PCR enzyme is able to PCR 1 fg of
target gene from genomic... 阅读全帖 |
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b******r
发帖数: 111 | 3 Thank for the message of you both.
"你怎么抽DNA的，测纯度没有，浓度怎么测的"
-->lysis buffer containing proteinase K digests mouse tail overnight at 65
degree. Phenol/chloroform isolates, ethanol precipitate. Dissolve in TE
buffer(pH7.5). 280/260 ratio=1.7-1.9, 浓度=0.2-0.5ng/uL
I tested 7 forward primers x 15 reverse primers. The totally primer pairs
are 105. All works well to PCR 1fg of plasmid which contain target gene, but
always don't work when template contains both 1fg of plasmid and 0.1ng of
mouse genomic ... 阅读全帖 |
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L*******r
发帖数: 5448 | 10 反证都要kick，丢了的话也就是1TD+1FG变成2TD而已 |
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A**d
发帖数: 13310 | 11 man, 19 points equal to 2td plus 1fg, much different
from 20points. And why risk giving opponents momentum
after a great drive? |
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A**d
发帖数: 13310 | 12 "meaningless"? What an idiot. Since when 11pt gap was "meaningless",
while 20pt lead was too "conservative"? Right, instead of making it
8pt+1fg, any head coach would be happy taking a 12pt "meaningful"
deficit.
I get the point: 20 point lead is too big too complex for Falcons
fans to comprehend. Even the 27:26 vs 29:26 scenario with 34 seconds
to go is till too complex for a typical Falcon fan. |
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b*****a
发帖数: 14583 | 19 我认为9个td+1fg+1safety应该够了 |
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Y*****g
发帖数: 1481 | 20 我鳄不错了，巴马08年SUGAR BOWL可是输了2个TD，我鳄这场1个TD+1FG。要是稳扎稳打
，这球能追回来。 |
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b*****a
发帖数: 14583 | 21 应该按让分算
让1fg以内算46开
让1td以内算37开
2td算28开
2td+算19开 |
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