f***h
发帖数: 52 | 1 【 以下文字转载自 Family 讨论区 】
发信人: fisch (-_-), 信区: Family
标 题: 原来说某某省人怎么样是有根据的
发信站: BBS 未名空间站 (Sat Apr 21 13:26:43 2007)
我以前还以为是无稽之谈。刚知道有一个学科叫地域文化(regional culture - a
subculture). 这个学科可以解释一个地区的人的主要品质，口音等等，并且这些都跟
当地的自然生态相关。看来是个很深刻的学科。
btw
据说南京人主要是河南移民 ~~~>_<~~~ |
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C******8
发帖数: 602 | 2 最近一段时间长这个cell总会发现media里脏乎乎的，有一些不知道是deris还是啥，
每次多用1xpbs多洗两遍也不行。。。
倒应该真不是mycoplasma contamination，因为前几天测还是negative的。。。
和以前的subculture也没多大区别啊。。。1200rpm5分钟centrifuge的 |
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D****g
发帖数: 275 | 3 当然能Proliferate，否则怎么Subculture培养的． |
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D****g
发帖数: 275 | 4 it proliferates very fast, you have to subculture the cells every 2-3 days. |
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R*****o
发帖数: 14902 | 5 请问subculture用多少tranfer, 10%?
. |
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c******r
发帖数: 3778 | 6 呵呵，没想到在这里看到你啊
http://www.atcc.org/ATCCAdvancedCatalogSearch/ProductDetails/ta
这个cell line当然可以proliferate。但是应该不是长特别快的，doubling time应该
在24小时以上。因为上面atcc的网站上说1：3-1：6的subculture ratio，2-3天需要
split，那么差不多一天左右一个doubling，这个是多数tumor cell line的正常
doubling time。
然后也不需要什么特殊的growth factor，就是加点serum就可以了。应该算最容易的
cell line啦。
good luck！ |
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s******s
发帖数: 13035 | 7 那个啥，目的就是要挑抗性克隆么？不能直接dilute在96孔板么？为啥还要P1/P2/P3?
1 |
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s*********r
发帖数: 22 | 8 我最近刚做的stable cell line , 用的是pCDNA3.1 载体
用的是zeocin筛选，不用G418.开始时候我自己用G418筛选， 老板告诉我是错的，要用
Zeocin筛选。
我尝试过转染293和293HK。
转染是regular钙转。 细胞密度是60%。转染后6小时就加药（也尝试过12小时）
zeoxin浓度是400ng/ml
基本上等conflent后，大概3-4天，1：2到2个100mm dish，5天后很多细胞死了，此时
转移到60mm dish，等single coloney . 然后等2周 ，看到很多coloney长出来， 然后
pick up到48-wells .
1 |
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w********r
发帖数: 1431 | 10 听你的意思是要收mix population,而不是single colony
我的经验是，这种混起来的stable cell通常表达很差，
最好挑好多单个的克隆，然后选几个表达高的，然后每次实验的时候两三个Stable
line 一起用。
我们的策略是在24孔板里做transfection,然后分到6cm开始加药杀，再把细胞分到若干
10cm的盘子里继续杀，关键是密度不能高，要保证最后可以挑到单克隆。调出来的单克
隆放到24孔板里慢慢养，同时做WB检测表达。表达好的就留下，表达不好的就扔掉。
1 |
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i***0
发帖数: 160 | 11 转染48h后，转到100 mm plate上，加药，每三天换一次液，一至两周后去掉药，等细
胞长满，分到新的100 mm 板上， 一千，500， 200 个细胞/板， 观察单克隆， 用灭
菌滤纸（3mm直径）占trypsin用灭过菌的镊子夹着在单克隆上一抹，然后把滤纸放入新
鲜培养液中（24孔板），immunostain，WB，来确认单克隆的表达量和表达均一性。 |
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n********k
发帖数: 2818 | 12 well,it is a problem-solving issue: there are always many choices and I
would think it over and over like with any complex experimentations: what
are the options and what might be a best one to approach this...As far as I
am thinking, the less passaging the better...otherwise one might be running
a risk of picking the cells from a same clone multiple times; as well, the
clonal growth/death bias over the passaging could also dramatically change
the properties of the culture over time...numerous ... 阅读全帖 |
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l**********1
发帖数: 5204 | 13 RE:
孵蛋的学第/妹 giantbird05 (大鸟)
Did you use Claycomb-medium ?
if not, please use it
Claycomb-medium (Sigma, Germany)
plus fibronectin/gelatine-precoated cell culture flasks.
more details about stuff protocol
pls go to
PLoS One. 2012; 7(2): e31669.
PuMeb link:
//www.ncbi.nlm.nih.gov/pmc/articles/PMC3285183/
citation:
>Cell culture
>The HL-1 cardiomyocytes (murine atrial tumor cell line) were
maintained in monolayer culture with Claycomb-medium (Sigma,
Germany), supplemented with 10% fetal calf serum... 阅读全帖 |
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l**********1
发帖数: 5204 | 14 注意media Lot的原产国
最重要的一个 可能是
去甲肾上腺素
1% norepinephrine (Sigma-
Aldrich, Germany)
from
北美的有的牛来源的原料
其中的疯牛病因子 and 瘦肉精因子等比德国的EU标准高
以下的所有东西 订货 按原产国来 订货 then try again.
Cell culture
>The HL-1 cardiomyocytes (murine atrial tumor cell line) were
maintained in monolayer culture with Claycomb-medium (Sigma,
Germany), supplemented with 10% fetal calf serum (SAFCBiosciences,
USA), 1% penicillin/streptomycin, 1% glutamax
(both Invitrogen, Germany) and 1% norepinephrine (Sigma-
Aldrich, Germany) in an incubato... 阅读全帖 |
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h******y
发帖数: 351 | 15 MEF would grow faster in 3% O2 than in 20% O2. Maybe you can try different
seeding and subculture schedule? |
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y********g
发帖数: 782 | 16 紧急求助，我的细胞是怎么了？
左边是换media 2小时候。右边是换media之前。
细胞是昨天上午subculture的，今天上午9点做实验，和treat细胞。 这个是Control
well的，就是只是change 了media. 然后下午1点就发现变成这样子。这种情况连续发
生2个星期了，开始以为药有问题，反复配了很多次，还是一样。后来有怀疑是media有
问题，又配了新的media. 现在这批，连细胞都是从liquid N2 搞得新的，还是这样子
，快崩溃了。。。。请大牛帮忙看一下~~~
换了media后，像是细胞长了个大水泡。。。 |
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w*****t
发帖数: 179 | 17 we are using the same system, but we didn't the the pRC plasmids, we have
our own different seratype AAV RC. I think it can be amplified, the same as
we used other RC plasmids. Also AAV-293 could be subcultured, but it is
better to use low passage cells to package the AAV. Good luck. |
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a******r
发帖数: 38 | 18 The conclusion of the argument could be understood as that a staggering
number of subcultures threatens the national identitie, which indicates a
conflicting relationship between the two concept. However, the answer choice
suggests the opposite, that national identity is not threatened by the
modern culture, but contributes to the development of it.
What do you say? |
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A*********r
发帖数: 8271 | 19 The assumption here is that national identities cannot be composed of
subcultures. Attacking the assumption can therefore weaken the conclusion.
great
off
the
commentator' |
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w*********g
发帖数: 82 | 20 Can you explain a little more about the assumption you discovered?
I didnt see any nexus between subcultures and fashions, ideologies,
religions, artistic movements, economic theories,cults, and dogmas in
fabulous profusion.
The only word makes me feel that is the right answer is "staggering". That
makes the statement as a whole incredible and less convincing.
Please help. Thank you. |
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D**********s
发帖数: 320 | 21 fashions, ideologies, religions, artistic movements, economic theories,
cults, and dogmas = subculture
spin off = create/produce here |
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l**********1
发帖数: 5204 | 22 PMID papers:
a,
Jozifkova E et al.
Evolutional background of dominance/submissivity in sex and bondage: the two
strategies? (2012).
Neuro Endocrinol Lett. 33:636-42.
CONCLUSION:
The preference for sexual arousal by dominance and submission may be
connected to strategy respecting rank, while the preference for "bondage"
may be derived from an opportunistic strategy that may be essential for
possible partner problems solution. From the evolutionary biology point of
view, these patterns of sadomas... 阅读全帖 |
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j****e
发帖数: 185 | 23 the resurrection
ALMOND: After the whole scene died down, it found its way into the fetish
scene, because the clubs kept playing gothic and industrial music. There are
clubs like the Torture Garden in London which is predominantly fetish, but
it's very gothic as well.
BERGER: Goth pretty much died out in the U.K. in the early '90s. It goes
through cycles in the U.S.-it dies, gets reborn over and over again. It's
probably died about five times now. Being that it's a vampire subculture, the
idea o |
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