w******n
发帖数: 767 | 1 没啥讲究吧，只不过用不着random了。我有时检测多个gene，用oligo(dt),做完反转录
，取RT产物可以检多个gene，没有问题。
primer |
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h*******o
发帖数: 4884 | 2 没有完全看懂楼主的意思 , 个人理解
所有的One step RT-PCR kit都不可能用gene specific primer来做RT这一步
都是用的oligodT之类的primer来做RT,然后在接着标准的PCR而已
只不过对Taq的要求高一点而已,需要HotStart Taq
user自己加入自己的specific primer for gene of interest.
整个mixture其实就是先RT在PCR, 一般Protocol都要求42度1小时RT,然后直接65赌灭活,再开始PCR的cycle.
我以前用过很多One-step RT PCR,如果不出条带一般是自己的引物的问题
至于One-step realtime PCR 没试过
primer |
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l**********1
发帖数: 5204 | 3 OMG
LZ used degenerated primer for cDNA single strand synthesis
R U Seriously? an amateur molecular biologist LZ?
Please go to
//www.protocol-
online.org/prot/Molecular_Biology/RNA/Reverse_Transcription__RT____cDNA_Synthesis/index.html
RT reaction is also called first strand cDNA synthesis. Three types of primers can be used for RT reaction:
oligo (dT) primers, random (hexamer) primers and gene specific primers with each having its pros and cons.
or
/media.affymetrix.com/support/technical/usb... 阅读全帖 |
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g**********y
发帖数: 423 | 4 GSEA can be applied as long as you have a ranked gene list, not limited to
gene expression.
I have used it for mutation analysis. |
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l**********1
发帖数: 5204 | 5 if by GFP not luciferase
you can try CAG (cytomegalovirus (CMV) early enhancer element and chicken
beta-actin) promoter
paper link:
//www.ncbi.nlm.nih.gov/pubmed/22378886
Mice strain link:
//jaxmice.jax.org/strain/016532.html
Description
The CAGs-rtTA3 transgene has a CAG (cytomegalovirus (CMV) early enhancer
element and chicken beta-actin) promoter driving widespread expression of
reverse tetracycline-controlled transactivator 3 (rtTA3). High rtTA3
expression is observed in most tissues examin... 阅读全帖 |
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z****u
发帖数: 1007 | 6 【 以下文字转载自 Neuroscience 讨论区 】
发信人: zhaohu (天外飞猪), 信区: Neuroscience
标 题: 用哪个cre 能特异knockout 某个gene 在periphery sensory nerv
发信站: BBS 未名空间站 (Sat Apr 14 03:11:06 2012, 美东)
不是做神经的，最近发现某个颌面部的干细胞分布和peripheral sensory nerve fiber
紧密相关。想找个inducible cre来在sensory nerve fiber里knockout掉一些gene.
请教该用啥cre比较好啊。最好是inducible cre. 多谢先了。 |
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a*****s
发帖数: 131 | 7 Hello everyone, please help: how to detect low abundance gene expression?
Now I use 1 ug of RNA per 20 ul cDNA reaction, then I use 1 ul of the cDNA.
For beta-Actin, the qPCR picks up the signal at around 15 cycles in linear
range. But my target gene (I used 2.5 ul per reaction) can only be picked up
at around 35 cycles (1 or 2 out of triplicated reactions), where it is
already out of the linear range!! How to enhance the signal?? Please help!
Thank you very much!! So frustrated... |
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l***y
发帖数: 638 | 8 pre-amp your sample for 10-15 cycles, that will bring down the Ct of your
gene to 20ish, but you might need to find a different housekeeping gene to
normalize to because Ct of actin will be below 10.
.
up |
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h***e
发帖数: 232 | 9 Many people normalize the expression to the housekeeping genes expression;
but what if I don't know how many normal/cancer cells, considering both
normal/cancer cell contribute to the housekeeping gene expression?
heterogeneous? |
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l**********1
发帖数: 5204 | 11 So,
better is to perform multiple GSA running,
pls refer attached figure
cited from
PMID 23444143
Väremo L et al., (2013).
Enriching the gene set analysis of genome-wide data by incorporating
directionality of gene expression and combining statistical hypotheses and
methods.
Nucleic Acids Res. 41: 4378-91.
>http://www.ncbi.nlm.nih.gov/pubmed/23444143 |
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s******a
发帖数: 252 | 12 I have written reviews on pathway analysis. Pathway, GO and gene sets
analyses are
just bunch of statistical methods. Like any statistical method, it depends
on how you use it and interpret it. Scientific publications often demand
evidence beyond a single statistical method.
One should consider both the quality of gene sets and the underlying
statistical method. GO is often less specific than pathway analysis. MsigDB
(database behind GSEA) is getting too big - one should be careful too. The
type... 阅读全帖 |
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h*********r
发帖数: 2844 | 13 向这里的生物大牛讨教一下：
股票班上有人看好pacbio， 有人bash。看了一圈，觉得PacBio 技术很多拿来做gene
full assembly， 很多和ilumina 测序hybrid。 这种科研除了发paper，养活postdoc
外，有多少实际用处？ 如果第一人做出来，还算有点创新，后人跟上，各种gene 都去
重做一遍，不是炒冷饭浪费钱吗？NIH一砍钱，不就砍这种项目吗？
当然要找科研意义，都说能挖出来的。funding 总是写的天花乱坠，我想问的是大方向
上的用途，尤其是能用于商业开发的价值。 |
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S*********e
发帖数: 127 | 14 a mutant has been mapped to a gene ~9kb. need to do rescue experiment to
confirm the gene specificity of phenotype. RNA injection apparently not
doable since its easy degradation. DNA injection mosaicism is not penetrated
enough to recover the whole phenotype. any suggestions? |
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s*****i
发帖数: 315 | 15 Gene越大包装效率越低，你给自己的gene加个小tag，感染完了染色看看吧
没做过in vivo，不太清楚。
体外做时我从来不提纯病毒的，直接用medium感染，但是效果很好
你的靶细胞基本不会分裂，即使感染成功，整合到基因组的效率不会太高的。不知道有
没有基于transposon的系统
★ 发自iPhone App: ChineseWeb 8.7 |
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Z******t
发帖数: 206 | 16 Genes Dev IF=10
Nat. Cell biol IF=19
两片Genes Development 找工作是不是上不了short list?
一片Nature Cell biology 是不是就可以？ |
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m******5
发帖数: 1383 | 17 弱问，现在在用
cufflink-cuffmerge-cuffdiff-cummeRbund这套流程分析手上的一些RNA-Seq data
cummeRbund的很多function和plot output都是cuffID，怎么把cuffID转成gene symble?
或者有什么R package可以batch retrieve gene symbol from cuff ID么? |
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M******g
发帖数: 152 | 18 关键字: protein categorization (gene ontology (GO) analysis
发信站: BBS 未名空间站 (Wed Apr 6 13:49:28 2016, 美东)
Can anybody recommend a good software or server to do protein categorization
(gene ontology (GO) analysis)?
Thank you. |
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G******n
发帖数: 289 | 19 你GO term给的gene list 应该是你考虑过gene length之后算出来的一个list， 所以
在GO的时候不用考虑length
TF的话，要看你用什么办法算的，如果是简单的像DAVID一样的enrichment，那也不用
考虑。其他的model需要考虑 |
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发帖数: 1 | 21 【 以下文字转载自 Postdoc 讨论区 】
发信人: tonytonyvan (), 信区: Postdoc
标 题: 实验求助：gene rescue
发信站: BBS 未名空间站 (Tue Aug 16 21:13:59 2016, 美东)
最近在做gene rescue，试了一次了，不太成功，但不知道问题出在哪里？不知道哪位
大师做过，请指点。 |
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c*****t
发帖数: 520 | 22 【 以下文字转载自 Biology 讨论区 】
发信人: cityant (cityant), 信区: Biology
标 题: Mathematical Models of Gene Regulation and Neuroscience.
发信站: BBS 未名空间站 (Sun Mar 30 11:03:50 2008)
请问各位，现在研究mathematical models for the dynamics of gene regulatory
networks的情况如何呢？是一些数学工作者在自娱自乐，还是这个领域还有点应用的前
景？另外听说neuroscience在生物研究里不太热了。不知computional neuroscience的
前景如何？
似乎越来越多做数学的人在做这些，而且不少人认为这些方向很容易拿教职。前几周一
个生物数学圈子里还可以的美国本土教授在演讲中说：“（大意）生物数学的状况不好
，数学界的人不知道我们在做什么，生物界的人看不起我们，只是这行好拿tenure
track。”我很不解。难道生物数学这个行当没有生物学的竞争激烈？按说如果这些
models |
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J********n
发帖数: 534 | 23 【 以下文字转载自 SanFrancisco 讨论区 】
发信人: JohnnyChan (陳強尼), 信区: SanFrancisco
标 题: 轮回，从GENE到DNA
发信站: BBS 未名空间站 (Thu Mar 12 13:18:53 2009)
October 14, 1980
Initial Public Offering (IPO) 1,000,000 shares offered at $35.00
Genentech stock trades on NASDAQ under symbol GENE
March 2, 1988
Genentech stock begins trading on the NYSE under the symbol GNE
October 25, 1995
Genentech stockholders approve Roche's extended buyout option Each share of
Genentech Redeemable Common Stock (CUSIP 368 710 208) is exchanged |
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P*****s
发帖数: 375 | 24 well, I'm a newer at bichemistry, so just discuss this rather than give
an answer.
I think they just define endonuleases as those -ases that can cleave
DNA at target sites. This is a must for the insertion of intron. So
there seems no way to clone gene without it. (Or you may ask: Can a
gene be cloned without cleaving DNA?) |
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c*****t
发帖数: 1879 | 25 Easy, I will just point a region on a bacterial chromosome and
say that's a gene. Unless you could give a precise definition of
gene cloning in 1950s, I wouldn't even bother. |
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p********a
发帖数: 5352 | 26 ☆─────────────────────────────────────☆
vincentsg (vincentsg) 于 (Tue Feb 5 11:42:22 2008) 提到:
i have 2 genes ---A and B , A 有 10 SNPs , B有6 SNPs.
A的10个SNPs中， 有9个 与某病X有关系， ( i did the model X~SNP for each SNP
separately)
B的6个SNPs中， 有0个 与某病X有关系， ( i did the model X~SNP for each SNP
separately)
怎样fit 1 个 model to show effect of GENE A as 9 of 10 SNPs are relatted
with
某病X?
what is the sas code?
Thanks!
☆─────────────────────────────────────☆
sir ( 郎 ) 于 (Tue Feb 5 13:07:38 2008) 提到: |
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h******1
发帖数: 94 | 27 In the manual,
http://www.broadinstitute.org/gsea/doc/GSEAUserGuide.pdf
On the page 7, right above "Phenotype Labels", it says
"The GSEA algorithm does not filter the expression dataset and does not
benefit from your filtering of the expression dataset. During the analysis,
genes that are poorly expressed or that have low variance across the dataset
populate the middle of the ranked gene list and the use of a weighted
statistic ensures that they do not contribute to a positive enrichmen |
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f****h
发帖数: 41 | 28 【 以下文字转载自 Biology 讨论区 】
发信人: futsch (MAP), 信区: Biology
标 题: suggestion for gene synthesis
发信站: BBS 未名空间站 (Wed May 18 18:22:32 2011, 美东)
Dear all,
I would like to synthesize a bacteria gene in a codon-optimized version for
the expression in mouse neurons.
Can anyone share some experience regarding this service from different
companies, like IDT, DNA2.0,
GenScript? Thank you! |
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z*******n
发帖数: 1034 | 29 Sorry, folks: Study says musical talent mostly comes from your genes
BY Jon Fingas @jonfingas 8 hours ago 48
We like to imagine that musical talent is just a matter of putting in enough
hours. The Beatles became superstars because they spent years honing their
craft in Hamburg, right? Well, maybe not. A recently published study from
the Karolinska Institute's Miriam Mosing suggests that you need the right
genes to become a true maestro. The research compared thousands of identical
and fraternal ... 阅读全帖 |
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d*****u
发帖数: 17243 | 30 强奸犯、杀人犯这些都跟gene有correlation的 |
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B******g
发帖数: 89 | 31 不信的可以搜索bacterial gene swapping.
不同种类的细菌可以交换一些有用的dna片段,互通有无.
现在发现一些鱼类也可以.
大力推广一妻多夫,可以大大加快人类进化的速度. |
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x**8
发帖数: 1939 | 32 搜索结果表明Gene Haddlebury这个人不存在，
相关新闻都是the onion杂志愚弄嘲讽大众的愚蠢， |
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n*****8
发帖数: 19630 | 33 那你是鸵鸟。
:-)
: 搜索结果表明Gene Haddlebury这个人不存在，
: 相关新闻都是the onion杂志愚弄嘲讽大众的愚蠢，
|
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y*j
发帖数: 3139 | 34 又一个张拾迈教授。
：搜索结果表明Gene Haddlebury这个人不存在，
：相关新闻都是the onion杂志愚弄嘲讽大众的愚蠢， |
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g**y
发帖数: 366 | 35 最近开始准备攒审稿，广发邮件几十封，有零星几个回复说把我加到reviewer list，
但是两个月了一篇也没收到，真急阿。
恳请这里的好心人若有合适的稿件考虑一下我，或者请帮我推荐一下，非常感谢！
我的研究领域是：computational biology, gene regulation, epigenetics, Next-
generation sequencing, microarray data analysis, machine learning, data
mining, clustering/classification etc.
自己发表过几篇computational biology, algorithms 等方面的paper, 最高impact
factor 8.02
若需要详细个人信息、publication list的话，请站内联系，谢谢！ |
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l******2
发帖数: 2994 | 37 "Our genes provide us with dispositions and tendencies toward personality
characteristics, and our experiences shape us throughout our lifetime. In
the early childhood years, children develop a core personality and sense of
themselves. They develop a view of the social and physical world and their
abilities to navigate the currents and shoals that carry them along" |
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l******2
发帖数: 2994 | 38 I feel most guys that followed the LZ focused one and ignored another. but
these two things work together to shape the future of a kid. For most of
people, genes don’t mean doomed, the environment have ample space do the
modification job. Also LZ wants to tell the parents that for all the
perspectives (not just the language, IQ, but more hidden culture or social
things ) it is better to send kids to the daycare. Some parents obliviously
only focus on the things that they can measure such as lang |
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J******o
发帖数: 6 | 39 杂志 Drug Target Insights is looking for 4 个 guest editor, 每个editor 需要
征集5篇PAPER， topics 是关于 Gene Therapy and Proteomics. 有兴趣的junior
scientists 留下email. 对申请EB1A有帮助。我是lead guest editor, 也是为了申请
绿卡用才做的。大家互相帮忙. |
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发帖数: 1 | 40 最近在做gene rescue，试了一次了，不太成功，但不知道问题出在哪里？不知道哪位
大师做过，请指点。 |
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L**********n
发帖数: 87 | 41 【 以下文字转载自 xiaoyaogu 俱乐部 】
发信人: LisaHannigan (起舞弄清影，曾照彩云归。), 信区: xiaoyaogu
标 题: SGMO ZFN GENE THERAPY to cure HIV
发信站: BBS 未名空间站 (Thu Mar 6 10:08:02 2014, 美东)
昨天5点NEJM， NYT出消息
盘后大涨30%以上（从19涨到24）
今早22-23入仓，看能不能近期翻番：） |
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x****9
发帖数: 2668 | 43 这必须顶一下。。。厉害。。。
不是针对gene。。。针对所有东东也是如此。。。不是吗？ |
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d***u
发帖数: 99 | 45 那你要捂 还是要割啊？
Genetic Technologies LtdNASDAQ:GENE*
*Nasdaq FSI: *Deficient: Issuer Failed to Meet NASDAQ Continued Listing
Requirements
搞不好就成垃圾股了。。。。 |
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n*****0
发帖数: 1721 | 49 这个gene我观察了很久了，每次涨起来都很厉害的样子，还都带着量
而且传说庄家早就跑光了啊，现在有两家基金买入什么的。。。 |
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w********8
发帖数: 665 | 50 敢问大牛gene还在手上没？ 这个帖子里有好几个资深大牛提了建议，100个包子发给谁
了。 |
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