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NGS 二代测序分析，大家来评评

NGS 二代测序分析，大家来评评# Biology - 生物学

M*o2011-10-03 07:10

1 楼

有时候五点多些去day care接娃，他已经累得有些犯困了。
于是到家就得赶紧给他喂饭，吃完饭后有时能玩一会儿，有时就困得不行了，得赶紧给
他洗漱准备睡觉。
大家说这样正常吗？多谢！

g*f2011-10-03 07:10

2 楼

父母即将于三月份来美国，她们第一次来。刚刚听一个同事说他前不久从国内回来在飞
机上什么表格都没填。我记得是要填一个海关申报单，还有个好像I94表什么的。请问
有最近来美国的吗？在飞机上需要填什么表格呢？我父母都不会英文，我想要填什么的
我先替他们填上，然后让他们在飞机上照抄就可以了。多谢

s*q2011-10-03 07:10

3 楼

去看了版上大热的Gone Girl和Interstellar，发现现在电影真的看不懂。看完Gone
Girl还是不明白女主角干嘛要这么做。觉得自己老公渣离婚不就好了，干嘛要搞的这么
复杂。还是说导演想展示精神不正常的夫妻生活是怎样的？

d*e2011-10-03 07:10

4 楼

听说和国内的重点班一样，刷题黄人比其他大厂多，内部竞争激烈，怎么网站还是一坨
屎样，界面和很多年前一样乱糟糟的，点开一个照片进度条都要闪几秒钟。

n*k2011-10-03 07:10

5 楼

Just got some of my ChIP-seq data back:
For histone markers, looked great and the cores told me it is great above 70
% mappable reads and peak calling with FDR of <5%...
for my TF, the data make sense to me but the core said it is trash/useless,
9-20% mappable reads (out of 9-11M, meant to get 20M) and peaks calling with
a FDR of 100%.
Luckily my TF has been chipped many many time and has very conserved binding
sites. I randomly picked the mapped peaks, most of them with at least 1
high confident binding site...so biology tells me the data is very good...
With that, I am thinking I would need to get a bit bioformatical myself...
So any guru on the field could make more comments on the data-mining/
analysis part...
Also, any recommendations on de nova motif finders from ChIP data? How
about duplicate reads, and repetitive regions?...Pretty frustrated as of now
,...either I haven't found the right places/softwares, or the data-mining/
analysis for NGS is still pretty much at a primitive stage...

M*o2011-10-03 07:10

6 楼

补充：他在day care一般是中午１２点到１点多不到２点午睡。

h*s2011-10-03 07:10

7 楼

只用海关报关单。 i94电子化了，不用自己填
[在 griefleaf (griefleaf) 的大作中提到：]
：父母即将于三月份来美国，她们第一次来。刚刚听一个同事说他前不久从国内回来在
飞机上什么表格都没填。我记得是要填一个海关申报单，还有个好像I94表什么的。请问
：有最近来美国的吗？在飞机上需要填什么表格呢？我父母都不会英文，我想要填什么
的我先替他们填上，然后让他们在飞机上照抄就可以了。多谢
：...........

r*e2011-10-03 07:10

8 楼

家版神经病多着呢

【在 s*******q 的大作中提到】

: 去看了版上大热的Gone Girl和Interstellar，发现现在电影真的看不懂。看完Gone
: Girl还是不明白女主角干嘛要这么做。觉得自己老公渣离婚不就好了，干嘛要搞的这么
: 复杂。还是说导演想展示精神不正常的夫妻生活是怎样的？

a*g2011-10-03 07:10

9 楼

Your core is right. Normally, 70% mappable reads are mandatory for a good
ChIP-seq. 20% is way too low to convince people this ChIP-seq is valid.

70
,
with
binding

【在 n********k 的大作中提到】

: Just got some of my ChIP-seq data back:
: For histone markers, looked great and the cores told me it is great above 70
: % mappable reads and peak calling with FDR of <5%...
: for my TF, the data make sense to me but the core said it is trash/useless,
: 9-20% mappable reads (out of 9-11M, meant to get 20M) and peaks calling with
: a FDR of 100%.
: Luckily my TF has been chipped many many time and has very conserved binding
: sites. I randomly picked the mapped peaks, most of them with at least 1
: high confident binding site...so biology tells me the data is very good...
: With that, I am thinking I would need to get a bit bioformatical myself...

o*22011-10-03 07:10

10 楼

我家的4岁，放学路上就能睡着，勉强吃完饭，洗漱，玩一会就睡了。我估计他中午不
睡眠。尽管老师
写的睡了。正常吧。

【在 M****o 的大作中提到】

: 补充：他在day care一般是中午１２点到１点多不到２点午睡。

g*f2011-10-03 07:10

11 楼

多谢了。还跟同事争论了半天呢，怎么会什么表格都不填。

s*i2011-10-03 07:10

12 楼

婚上十年以后就看懂了。

【在 s*******q 的大作中提到】

R*n2011-10-03 07:10

13 楼

If I want to do a methylation ChIP-seq on SOLiD or Illumina's system, how
many reads should I require from the Core. I asked a Third part company.
They mention that each run can fit 10 samples, about 6.6 GB data per sample.
I don't know how to select the best throughput for the methylation ChIP.
Thank you!

【在 a******g 的大作中提到】

: Your core is right. Normally, 70% mappable reads are mandatory for a good
: ChIP-seq. 20% is way too low to convince people this ChIP-seq is valid.
:
: 70
: ,
: with
: binding

M*o2011-10-03 07:10

14 楼

多谢回复！
他应该是中午真的午睡了。因为我有时趁着吃午饭时偷偷溜去看看他。他们toddler班
上所有的小朋友都能被老师给弄着了。呵呵。
我主要是觉得一来刚吃完晚饭就睡觉怕对肠胃不好，二来没啥太多时间和宝宝玩或亲近
。只能靠周末了。５５５

【在 o***2 的大作中提到】

: 我家的4岁，放学路上就能睡着，勉强吃完饭，洗漱，玩一会就睡了。我估计他中午不
: 睡眠。尽管老师
: 写的睡了。正常吧。

p*e2011-10-03 07:10

15 楼

跟导演有啥关系。。。

【在 s*******q 的大作中提到】

a*g2011-10-03 07:10

16 楼

I haven't done methylation ChIP-seq. For TF ChIP-seq on GAII from illumina,
20m should be fine.

sample.

【在 R****n 的大作中提到】

: If I want to do a methylation ChIP-seq on SOLiD or Illumina's system, how
: many reads should I require from the Core. I asked a Third part company.
: They mention that each run can fit 10 samples, about 6.6 GB data per sample.
: I don't know how to select the best throughput for the methylation ChIP.
: Thank you!

a*e2011-10-03 07:10

17 楼

拍拍。。。。

l*k2011-10-03 07:10

18 楼

看懂是看懂了，还是想说一句：这特么就是深井冰

【在 s**i 的大作中提到】

: 婚上十年以后就看懂了。

n*t2011-10-03 07:10

19 楼

With that awful mapping rate, it is difficult to justify the result.

M*o2011-10-03 07:10

20 楼

妮妮～～～
bo~~~

【在 a****e 的大作中提到】

: 拍拍。。。。

s*i2011-10-03 07:10

21 楼

源于生活，高于生活嘛。Amy和Nick的互动由爱生恨，又在废墟上苟且延续，多少婚姻
里都在上演，电影用更戏剧化的手段表现出来。
另外，Scott Peterson,李天一，还有李昌钰破的wood chipper碎尸案，真实生活其实
比创作还精彩诡异。

【在 l**k 的大作中提到】

: 看懂是看懂了，还是想说一句：这特么就是深井冰

s*s2011-10-03 07:10

22 楼

never done mapping myself, but FYI
1. histone marker和TF的Chip-seq protocol应该非常不一样，不知道你是不是用的一
个protocol
2. hiseq现在一条lane大概200m reads, 50bp的话，略微不到40G的data.

70
,
with
binding

【在 n********k 的大作中提到】

j*f2011-10-03 07:10

23 楼

刚吃饭就睡着挺好的。呵呵。多舒服啊！我最喜欢吃完就睡了～

多谢回复！
他应该是中午真的午睡了。因为我有时趁着吃午饭时偷偷溜去看看他。他们toddler班
上所有的小朋友都能被老师给弄着了。呵呵。
我主要是觉得一来刚吃完晚饭就睡觉怕对肠胃不好，二来没啥太多时间和宝宝玩或亲近
。只能靠周末了。５５５

【在 M****o 的大作中提到】

: 多谢回复！
: 他应该是中午真的午睡了。因为我有时趁着吃午饭时偷偷溜去看看他。他们toddler班
: 上所有的小朋友都能被老师给弄着了。呵呵。
: 我主要是觉得一来刚吃完晚饭就睡觉怕对肠胃不好，二来没啥太多时间和宝宝玩或亲近
: 。只能靠周末了。５５５

D*o2011-10-03 07:10

24 楼

故事还是可以看看，但是没评分那么好
有人说什么是David Fincher最好的片子
只能说他没怎么看过他的片子
这个在Lynch导过的片子里算below average的了，
当然主要还是故事一般般吧
电影版的desperate housewife

n*k2011-10-03 07:10

25 楼

I did the ChIP part and the core prepared the libraries and I believe it was
the same protocol for all samples. Apparently TF groups have much lower
amount of DNA to start with and I was wondering whether that would make the
background noise much big an issue and thus the percentage of the mappable
reads is very low. I have three conditions: the percentage of the mappable
reads is decreasing as the same way the starting amount of ChIP-DNA does. I
was wondering whether over-amplification could cause the problem?
Nonetheless, for the mapped peaks, most of them are correct ones while the
rest needs to be verified further.

【在 s******s 的大作中提到】

: never done mapping myself, but FYI
: 1. histone marker和TF的Chip-seq protocol应该非常不一样，不知道你是不是用的一
: 个protocol
: 2. hiseq现在一条lane大概200m reads, 50bp的话，略微不到40G的data.
:
: 70
: ,
: with
: binding

h*i2011-10-03 07:10

26 楼

太可爱了。都睡的东倒西歪。。。

【在 M****o 的大作中提到】

y*n2011-10-03 07:10

27 楼

林奇意外躺枪

【在 D**o 的大作中提到】

: 故事还是可以看看，但是没评分那么好
: 有人说什么是David Fincher最好的片子
: 只能说他没怎么看过他的片子
: 这个在Lynch导过的片子里算below average的了，
: 当然主要还是故事一般般吧
: 电影版的desperate housewife

n*k2011-10-03 07:10

28 楼

Sure, I would agree from statistics, however, Since the gene as you know
have very conserved binding site and have been ChIPed so many time----what
would be ur thoughts on the fact: most of the peaks (1200 for one condition
and 800 for a 2nd---the numbers also agree with my expectation between the
two conditions) are confirmed targets while the rest could be potentials
ones...

【在 a******g 的大作中提到】

: Your core is right. Normally, 70% mappable reads are mandatory for a good
: ChIP-seq. 20% is way too low to convince people this ChIP-seq is valid.
:
: 70
: ,
: with
: binding

vn2011-10-03 07:10

29 楼

第一次看到美国还有这样睡觉的床
为啥我们这边就见不到

g*t2011-10-03 07:10

30 楼

同意，我也觉得fincher很棒，把电影拍得很好，可是受故事限制，确实比不上他别的
片子。

【在 D**o 的大作中提到】

L*a2011-10-03 07:10

31 楼

Generally, 70% mappable reads is an experienced criteria.
FYI.
ENCODE project requires 10M mappable reads for human ChIP-seq experiments,
and modENCODE project requires 4M for worm and fly.
From the perspective of genomic, some classic genes bound by peaks are not
enough to prove your ChIP-seq data is valid, in terms of saturation.
You'd better to have some replicates anyway, each of replicate should meet
saturation and mappability criteria, and the reproducibility rate also
should be in an acceptable range (e.g. 80% overlapping)

V*82011-10-03 07:10

32 楼

sooooo cute

【在 M****o 的大作中提到】

r*e2011-10-03 07:10

33 楼

这片是给夫妻看的

【在 g********t 的大作中提到】

: 同意，我也觉得fincher很棒，把电影拍得很好，可是受故事限制，确实比不上他别的
: 片子。

j*p2011-10-03 07:10

34 楼

1. which species?
2. whether your TF has been chipseqed before?
3. does the core use MACS to call peaks for your TF?

70
,
with
binding

【在 n********k 的大作中提到】

M*o2011-10-03 07:10

35 楼

555
那就是个小垫子呀。哪里是床。。。
他们day care的infant班是睡crib的，但到了一岁以上的toddler班就是睡垫子了。
大家宝宝上的day care toddler class不是这样的吗？

【在 vn 的大作中提到】

: 第一次看到美国还有这样睡觉的床
: 为啥我们这边就见不到

g*t2011-10-03 07:10

36 楼

这意思是别人没看懂？

【在 r***e 的大作中提到】

: 这片是给夫妻看的

n*k2011-10-03 07:10

37 楼

rats
sure, many times
yes

【在 j*p 的大作中提到】

: 1. which species?
: 2. whether your TF has been chipseqed before?
: 3. does the core use MACS to call peaks for your TF?
:
: 70
: ,
: with
: binding

vn2011-10-03 07:10

38 楼

我觉得这个垫子比这边很多地方的cob都好呀 cob就是一个塑料网子小孩窝在里面肯定
不舒服对骨骼也不好吧不知道那垫子是不是全棉的呢？
反正这边睡觉的床土的不得了

【在 M****o 的大作中提到】

: 555
: 那就是个小垫子呀。哪里是床。。。
: 他们day care的infant班是睡crib的，但到了一岁以上的toddler班就是睡垫子了。
: 大家宝宝上的day care toddler class不是这样的吗？

D*o2011-10-03 07:10

39 楼

小说结尾和电影不一样，还打算拍续集？
小说里Amy回来后，还设计了一次计谋毒死Nick，同时计划写一本书
把自己做过的事情都坦白

【在 g********t 的大作中提到】

: 同意，我也觉得fincher很棒，把电影拍得很好，可是受故事限制，确实比不上他别的
: 片子。

n*k2011-10-03 07:10

40 楼

thanks for the infor, and was looking at the guideline...so, what would be
minimum number of cells for a TF-chip?

【在 L*******a 的大作中提到】

: Generally, 70% mappable reads is an experienced criteria.
: FYI.
: ENCODE project requires 10M mappable reads for human ChIP-seq experiments,
: and modENCODE project requires 4M for worm and fly.
: From the perspective of genomic, some classic genes bound by peaks are not
: enough to prove your ChIP-seq data is valid, in terms of saturation.
: You'd better to have some replicates anyway, each of replicate should meet
: saturation and mappability criteria, and the reproducibility rate also
: should be in an acceptable range (e.g. 80% overlapping)

vn2011-10-03 07:10

41 楼

还有真羡慕你家有个能睡的娃

j*p2011-10-03 07:10

42 楼

"for my TF, the data make sense to me but the core said it is trash/useless,
9-20% mappable reads (out of 9-11M, meant to get 20M) and peaks calling with
a FDR of 100%. "
Mouse sample with 20%x11M = 2.2M is useless for publication. But it is still
potentially useable for trouble shooting.
Possible reasons(most likely -- least likely):
1. Anti-body doesn't work, did not pull down anything, therefore, no signal
enrichment on sites that are supposed TF-binding. The whole signal should
look no difference between your input, you are supposed to see flat line (
except centermere) across chromosomes.
2. Library overload. this will give you less output reads, because 11M is
kind of low, usually GXII generates 20-30M reads, with raw .fastq file size
of 3-6G. If this is the only reason, you should still be able to see chip-
enriched sites, but fold enrichment should be low. you should upload your
mapped data to genome browser, or IGV and go to some of you positive
controls and take a look whether their promoters/enhancers have signal?
3. sequencing mapping, if raw reads is long, trim reads will give you
slightly better mapping, but won't change the mappable reads distribution.
which means if you did not see enrichment with 2.2M reads, you probably won'
t see enrichment in 4.4M reads.
"Luckily my TF has been chipped many many time and has very conserved
binding sites. "
If the same TF had been chipseqed many times, and if this TF has a conserved
motif, you can use it's motif to reverse search binding sites use fimo (
part of meme suite).
"I randomly picked the mapped peaks, most of them with at least 1high
confident binding site...so biology tells me the data is very good..."
This doesn't make sense to me, I never heard of something called "mapped
peaks", it should either be "mapped reads" or "called peaks". If it is "
mapped reads", of course if you blat them against genome reference, they'll
go to somewhere, but whether many of the reads will pile up together is
another question. I don't think this could be used as an evidence of showing
your data is very good.
"With that, I am thinking I would need to get a bit bioformatical myself...
So any guru on the field could make more comments on the data-mining/
analysis part...
Also, any recommendations on de nova motif finders from ChIP data? How
about duplicate reads, and repetitive regions?...Pretty frustrated as of now
,...either I haven't found the right places/softwares, or the data-mining/
analysis for NGS is still pretty much at a primitive stage..."
I wrote a brief introduction few days ago, only if you are interested.
Suggestion: tell your core to show you the mapped reads on browser, seeing
is believing, and take close look into those called peaks (I guess most of
them with FDR 100%), you'll then understand whether you should trust them.
BTW. the number of peaks means nothing, because one call always call more or
fewer peaks by manipulating thresholds.

g*92011-10-03 07:10

43 楼

睡得太少了，
从2点玩到5点够累了。

【在 M****o 的大作中提到】

: 补充：他在day care一般是中午１２点到１点多不到２点午睡。

j*p2011-10-03 07:10

44 楼

people use cell numbers from 10-100M to do normal TF chipseq. New
techniques are developing to chip in small amount of cells, as few as ~ten
thousands, as someone claimed. (for example: Single-tube linear DNADNADNA
amplification (LinDADA) for robust ChIP-seq)

【在 n********k 的大作中提到】

: thanks for the infor, and was looking at the guideline...so, what would be
: minimum number of cells for a TF-chip?

h*e2011-10-03 07:10

45 楼

哈哈，中间这个特写的就是你家娃吧，小朋友们真可爱啊。
大概睡着的时间不长吧。
他早晨几点醒呢？
我再深挖一下，你这也算BSO呢，还是特隐蔽那种，BSO娃早早就睡了你们可以做自己的
事了。

【在 M****o 的大作中提到】

n*k2011-10-03 07:10

46 楼

This is so great and thank you very much.
1. the Antibody is at least decent---this gene has been chipped many many
times by many labs; and I confirmed the ChIP using QPCR...
2. I have been suspecting we may have the library overload problem or over-
amplification issue(if that makes sense). The core really followed the
histone protocol and was meant to get 20-30M reads, and it did for Histone
markers and Input DNA. However, for my TF, the 1st is 9M with 9% mappable
reads (I expect less bind events than the second one) and the 2nd one is 11M
with 23% mappable reads--all after excluding duplicate reads.
3. Yes, it was called peaks-- many of the called peaks look very nice to me
using IGV software, showing very nice enrichment---the folds of the
enrichment range from 5-2000 across the genome (many are low). Among many I
have checked, the known binding site is sitting right at(or very close to)
the center of the peaks. That's what promoted me to think the data is still
usable but I might need to increase my starting materials in term of
saturation. I used the Solid/invitro new kit and it claimed to only need 10-
300K cells for ChIP-seq. I used about 300K for histone markers and half goes
to the library preparation, it worked nicely. I used about 2-4M for TF, and
the above is what I got...
4. Could you please check your mailbox. thanks.

useless,
with
still
signal

【在 j*p 的大作中提到】

: "for my TF, the data make sense to me but the core said it is trash/useless,
: 9-20% mappable reads (out of 9-11M, meant to get 20M) and peaks calling with
: a FDR of 100%. "
: Mouse sample with 20%x11M = 2.2M is useless for publication. But it is still
: potentially useable for trouble shooting.
: Possible reasons(most likely -- least likely):
: 1. Anti-body doesn't work, did not pull down anything, therefore, no signal
: enrichment on sites that are supposed TF-binding. The whole signal should
: look no difference between your input, you are supposed to see flat line (
: except centermere) across chromosomes.

M*o2011-10-03 07:10

47 楼

I've never seen the cob you mentioned. Even couldn't find a picture of it
by Google. hehe
Those pads my baby's day care uses have plastic cover. They are tri-folded.
:p

【在 vn 的大作中提到】

: 我觉得这个垫子比这边很多地方的cob都好呀 cob就是一个塑料网子小孩窝在里面肯定
: 不舒服对骨骼也不好吧不知道那垫子是不是全棉的呢？
: 反正这边睡觉的床土的不得了

n*k2011-10-03 07:10

48 楼

I was reading about the LinD, any experience or comments on it? thanks

【在 j*p 的大作中提到】

: people use cell numbers from 10-100M to do normal TF chipseq. New
: techniques are developing to chip in small amount of cells, as few as ~ten
: thousands, as someone claimed. (for example: Single-tube linear DNADNADNA
: amplification (LinDADA) for robust ChIP-seq)

M*o2011-10-03 07:10

49 楼

嗯。盖着小蓝被子的就是他。姥姥给亲手缝制的爱心牌小棉被子加个被罩。
他晚上有时会醒来哭几声。不理他的话就又睡过去了。早上七点多就醒。
BTW绝对没有BSO哈。我还发愁让他吃完晚饭怎么坐在potty chair上呢。因为他经常有
些constipated。基本每天都poo poo，有时隔天才poo poo。我们啥办法都试了，上
prune juice,dried prune,veggies and fruits, high fiber food, plenty of water
。可他有时poo poo还是很费力。甚至会在poo poo出来的时候哭一下子。很可怜。５５５

【在 h*********e 的大作中提到】

: 哈哈，中间这个特写的就是你家娃吧，小朋友们真可爱啊。
: 大概睡着的时间不长吧。
: 他早晨几点醒呢？
: 我再深挖一下，你这也算BSO呢，还是特隐蔽那种，BSO娃早早就睡了你们可以做自己的
: 事了。

m*c2011-10-03 07:10

50 楼

No matter how many reads you got from your core (raw data), only mapped
reads tell trues. From your mapper reads, you got the results that are in
line with your previous experiments. This tells you that the mapped reads in
your experiment makes sense. The minimum total number of mapped reads for a
chip seq shouldn't be a fixed number and it varies upon the genome and the
binding protein or histone markers that you are studying. Usually, histone
maker requires more mapped reads because histone marker simply distributes
more even and broadly. For some binding proteins, you may never be able to
get enough mapped reads as it is so unique and has no many binding sites.
With your mapped reads, calling for enriched genome region is going to be
important for you. I personally think the MACS will do fairly good work for
binding protein chip seq.
Good luck!

condition

【在 n********k 的大作中提到】

: Sure, I would agree from statistics, however, Since the gene as you know
: have very conserved binding site and have been ChIPed so many time----what
: would be ur thoughts on the fact: most of the peaks (1200 for one condition
: and 800 for a 2nd---the numbers also agree with my expectation between the
: two conditions) are confirmed targets while the rest could be potentials
: ones...

T*t2011-10-03 07:10

51 楼

正常。

【在 M****o 的大作中提到】

: 有时候五点多些去day care接娃，他已经累得有些犯困了。
: 于是到家就得赶紧给他喂饭，吃完饭后有时能玩一会儿，有时就困得不行了，得赶紧给
: 他洗漱准备睡觉。
: 大家说这样正常吗？多谢！

i*g2011-10-03 07:10

52 楼

never done the assay before, so i cannot tell

r*y2011-10-03 07:10

53 楼

5555,躲墙角哭去了，同是一岁半的小娃，我娃每天不到9点绝不肯睡，早上不到7点就
爬起来了。
他在daycare倒是睡很多，3个小时
poopoo问题也跟你的正相反，平均每天两次吧，刚上daycare老师经常没及时换，红屁
股。现在主管老师已经了解他了，可是一旦那个老师不在，还是会出问题

n*k2011-10-03 07:10

54 楼

thanks all the same for ur friendly response...

【在 i*****g 的大作中提到】

: never done the assay before, so i cannot tell

M*o2011-10-03 07:10

55 楼

呵呵，睡眠总数够了就行了吧。话说你娃他们day care每天都让睡那么长时间呀？那样
老师倒是省事了。呵呵。
我娃他们day care的老师都是每２－３小时换次diaper，我都觉得有些浪费了呢。:p
我们自己在家，或着尤其是带他出去玩，都不会换那么频繁呢。kaka

【在 r****y 的大作中提到】

: 5555,躲墙角哭去了，同是一岁半的小娃，我娃每天不到9点绝不肯睡，早上不到7点就
: 爬起来了。
: 他在daycare倒是睡很多，3个小时
: poopoo问题也跟你的正相反，平均每天两次吧，刚上daycare老师经常没及时换，红屁
: 股。现在主管老师已经了解他了，可是一旦那个老师不在，还是会出问题

b*r2011-10-03 07:10

56 楼

我是外行
不过FDR是啥东西我还是懂，是说的false discovery rate吧，peaks calling with a
FDR of 100%. 那不是说等于没有任何阳性结果？
另外你只有20%的序列可以map到genome里，其他都是些啥东西，你有没有看过，会不会
是什么东西污染，还是就是纯粹的noise？打个比方，如果是细菌或者其他实验残留的
污染，能不能找到污染源，减去这个background？这么大的序列应该能找到到底是啥东
西，如果不是noise的话

M*o2011-10-03 07:10

57 楼

给楼上的妈妈爸爸们都发包子了。谢谢大家哈！

j*p2011-10-03 07:10

58 楼

Agree.
Unmapped reads could be caused by (not limited to):
1. sequencing error. these reads probably won't map to any genome.
2. bacterial/viral contamination during library preparation. It won't be
easy to identify which contamination it is, if you don't have any candidates
ahead of time, however, if you do, it is pretty easy to confirm. We
recently found ~90% of our unmapped reads could be map to a bacterial genome
. This bacterial was used to replace bees to stick down the protein. while
in out input lanes, majority of the reads map to human genome.
Nevertheless, remove background won't help to increase local enrichment. and
the worse case is that you have a quite decent mappable rate, but they are
just randomly distributed to the entire genome, which looks no difference
with input.

a

【在 b****r 的大作中提到】

: 我是外行
: 不过FDR是啥东西我还是懂，是说的false discovery rate吧，peaks calling with a
: FDR of 100%. 那不是说等于没有任何阳性结果？
: 另外你只有20%的序列可以map到genome里，其他都是些啥东西，你有没有看过，会不会
: 是什么东西污染，还是就是纯粹的noise？打个比方，如果是细菌或者其他实验残留的
: 污染，能不能找到污染源，减去这个background？这么大的序列应该能找到到底是啥东
: 西，如果不是noise的话

r*y2011-10-03 07:10

59 楼

我娃的daycare也是2小时换一次的，但是娃皮肤敏感，poopoo完半小时内不换肯定出问
题，15分钟内不换好都会有一点点红

【在 M****o 的大作中提到】

: 呵呵，睡眠总数够了就行了吧。话说你娃他们day care每天都让睡那么长时间呀？那样
: 老师倒是省事了。呵呵。
: 我娃他们day care的老师都是每２－３小时换次diaper，我都觉得有些浪费了呢。:p
: 我们自己在家，或着尤其是带他出去玩，都不会换那么频繁呢。kaka

b*r2011-10-03 07:10

60 楼

我觉得noise如果来自某一个或几个生物来源还是很容易找的，拼起一小段到ncbi里去
blast一下不就出来了，什么genome和vector它都会去比较到的

candidates
genome
and

【在 j*p 的大作中提到】

: Agree.
: Unmapped reads could be caused by (not limited to):
: 1. sequencing error. these reads probably won't map to any genome.
: 2. bacterial/viral contamination during library preparation. It won't be
: easy to identify which contamination it is, if you don't have any candidates
: ahead of time, however, if you do, it is pretty easy to confirm. We
: recently found ~90% of our unmapped reads could be map to a bacterial genome
: . This bacterial was used to replace bees to stick down the protein. while
: in out input lanes, majority of the reads map to human genome.
: Nevertheless, remove background won't help to increase local enrichment. and

vn2011-10-03 07:10

61 楼

sorry... just found the name is cot...
thanks for the baozi!

folded.

【在 M****o 的大作中提到】

: I've never seen the cob you mentioned. Even couldn't find a picture of it
: by Google. hehe
: Those pads my baby's day care uses have plastic cover. They are tri-folded.
: :p

a*e2011-10-03 07:10

62 楼

抱抱，谢谢包子
试试吃橙子？
我家娃拉臭臭的时候我一般会给他揉一下肚子左后侧靠近肠子的位置
人家小肚子上肌肉不够强劲所以要使好大劲才行
还有你看看他屁屁上会不会有撕裂？

water
５５

【在 M****o 的大作中提到】

: 嗯。盖着小蓝被子的就是他。姥姥给亲手缝制的爱心牌小棉被子加个被罩。
: 他晚上有时会醒来哭几声。不理他的话就又睡过去了。早上七点多就醒。
: BTW绝对没有BSO哈。我还发愁让他吃完晚饭怎么坐在potty chair上呢。因为他经常有
: 些constipated。基本每天都poo poo，有时隔天才poo poo。我们啥办法都试了，上
: prune juice,dried prune,veggies and fruits, high fiber food, plenty of water
: 。可他有时poo poo还是很费力。甚至会在poo poo出来的时候哭一下子。很可怜。５５５

I*s2011-10-03 07:10

63 楼

太可爱了！另外，daycare一天玩得真的很累的，睡着太正常了。我们从5，6个月就这
样了，以前是在车上就睡着了，现在大了不至于，但是反正5点多回到家先要一个1小时
的nap，然后吃饭，玩会儿，再睡。

【在 M****o 的大作中提到】