c*m
发帖数: 1114 | 1 这两个玩意都是acronis出的，
true image专门clone/backup系统的。
director suite专门做disk partition/operation的。
这两玩意我都在用，人家说的没啥错，虽然director suite也能copy partition,但tru
e image要专业/强大的多。 |
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D**C
发帖数: 6754 | 2 我现在笔记本上83/256G
如果我要买128 GB m4，
1，可以吗？
2，如何才可以把83G的东西直接clone过来，我里面有不少数据，不想删掉
谢 |
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D**C
发帖数: 6754 | 3 我现在笔记本上83/256G
如果我要买128 GB m4，
1，可以吗？
2，如何才可以把83G的东西直接clone过来，我里面有不少数据，不想删掉
谢 |
|
s********a
发帖数: 1680 | 4 就是说有两个一模一样的笔记本，出厂win8.1系统，有一台的硬盘挂了，如果用另外一
台的系统clone到一个新硬盘上，然后装到这台挂掉硬盘的电脑上，进入系统以后，
windows能自动激活么。
也就是说笔记本在出场的时候，往硬盘上灌的image都是identical的么？含不含
license
相关信息？（我知道slic在bios里面） |
|
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U*****n
发帖数: 33 | 6 Thinkpad 450s reported SSD error. Got a Samsung 850 Pro and tried to make a
clone out of the old SSD. The Samsung software Data Migration got stuck at
11%. There might be bad sectors. Then searched and found some free software
called EaseUS, but it could not find the new SSD (connected with laptop with
an adapter).
Any one in a similar situation? Any suggestion is appreciated! |
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F***Q
发帖数: 6599 | 7
I've cloned over a dozen drives between computers (large drive to small,
small to large), always use a Ubuntu live USB, and the dd/pv command, never
had any issue.
for bad sectors, you need to use disk scan software to label those sectors
as "bad", then OS will not attempt to read from those locations.
If you use Ubuntu live usb, install a program called "badblocks" and use it
to scan your drive. I am sure you can find something similar for windows. |
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B******y
发帖数: 9065 | 8 像这种传统HDD往SSD里面clone系统的，需要看SSD而定，Samsung有自己的Data
Migration Software，WD有Acronis True Image WD Edition，去它们的官网下载就是
了。
貌似市场上没有见到过Seagate的SSD。 |
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g*****g
发帖数: 34805 | 9 Folks, I am confused when one needs to implement clone interface
in java, can't java just copy memory like C++? |
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c*****t
发帖数: 1879 | 10 int[][] abc = int[3][5];
int[][] def = (int[][])abc.clone ();
My question is that does def contains the reference of rows of abcs
or it's a complete duplication? It seems to me the first one is the
case, but want to make sure. |
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s********s
发帖数: 4011 | 11 很久没折腾，脱离现实了。上次克隆硬盘还是用的ghost7，请问现在
比较容易搞到又可靠的clone硬盘软件有什么？
还想把一些mkv的video转成avi好用dvd机放，有什么免费软件吗？先谢了。 |
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H*****i
发帖数: 1867 | 12 You misunderstand his poster.
Ghost 9.0 is very bad to clone system as image file. I never succeed. so
, please try acronis. acronis is very good.
there |
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g*******t
发帖数: 1039 | 13 thanks for the reply - but your conclusion is kind of suspecious (not to be
personal) - seems many people cloned system drive successfully in XP.
Do you know any reason that XP would log you off immediately while loading
personal seetings? (virus excluded) |
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y****e
发帖数: 71 | 14 If contamination goes 99.9% in all clone, that means you get a supercoiled
circular DNA as your contaminant. It transforms at a much higher efficiency.
First thing you might do is change plasmid.
I talked it to my boss, he had an interesting theory. If your insert confers
toxicity to E.coli, he said there might be some wierd recombination event that
cut the insert out of the plasmid, along with flanking reagions (which might
contain the MCS, that is why you cannot cut it again with your REs). |
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a******n
发帖数: 96 | 15 this is happening at hollywood, florida, but it is not a hollywood
story. a group, clonaid, which believes that life on earth was
created by et's, is to make the annoucement friday 9am est that
the birth of a cloned baby girl is imminent. |
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g***m
发帖数: 465 | 16 check NEB catalog for Taq enzyme, it has double strand specific 5'->3'
exonuclease activity.
My guess is primer and/or product degradation by Taq.
BTW, one small tip for pfx is: using 2X buffer rather than 1X. I got several
cDNA clones by PCR, and none of them disrupt the ORF by this modification.
DNA(WASP)
然后用同样的primer
colony-pGEX4t1
这些在处理后的pGEX4t1 |
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g****r
发帖数: 14 | 17 好处：efficiency, reliability
困难：you have to careful if you want to have some customized things
expensive, if your lab buy the system
I don't think there will be any issue for puslbishing, it is just cloning .... |
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r******m
发帖数: 173 | 18 I am trying to clone a 300 bp gene into pTYB2 vector (7.4kb)for later
protein purification method by IMPACT-CN. NdeI (producing sticky end) and
SmaI (blunt end) restriction sites were constructed into primers
corresponding to the N- and C- termini of the gene and incorporated by PCR
amplification. The PCR product and the vector were both double digested with
SmaI and NdeI respectively. After gel purification, the digested insert (
gene) and the vector were used to perform ligation with T4 DNA li |
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d*****r
发帖数: 2583 | 19 ☆─────────────────────────────────────☆
haoyun (福娃) 于 (Thu Dec 18 17:35:33 2008) 提到:
要抽100kb左右的BAC DNA用于end sequencing，哪个公司的kit比较好？
谢谢指教！
实验室里普通的plasmid我们都用Qiagen的midi prep kit
也可以用于BAC DNA extraction吗？
我以前用土法抽过BAC clone，恶梦啊
100ml culture然后用Qiagen midi kit够不够sequencing？
我以前土法抽BAC累不说，sequencing结果非常不好
☆─────────────────────────────────────☆
longwoodwalk (想做点啥) 于 (Thu Dec 18 17:41:14 2008) 提到:
用qiagen的high-speed maxi prep kit. 效果非常好。唯一注意的是，在elute你的DNA
之前，将Elution buffer (QF?) preheat到65-7 |
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t******r
发帖数: 8600 | 20 Invitrogen has some good stuff, like Topo, etc. But, most of its products
have very poor quality, I guess they
are getting too big and there are tons of garbage startups sequeeze their
poor products onto their shelf.
BAD!!!
The bad stuff I have used recently:
Stbl2 & 6
DNA ligase
Cre recombinase
RE enzymes
...
...
For cloning, NEB products are the most reliable. |
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A**E
发帖数: 191 | 21 For enzymes NEB is always the first choice.
However, Invitrogen's TOPO cloning kit is very convenient indeed. And
efficiency is quite high in my hands. |
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t****p
发帖数: 1504 | 22 topo cloning kit两年前还很好用。我们实验室有半年左右克隆出现问题，后来我买了
promega，qiagen的做对比，invitrogen的topo相比之下一塌糊涂。
坚决与invitrogen交涉，询问有没有人反映类似的问题，invitrogen断然否认，反复几
次之后我也放弃了。两个月前，隔壁实验室也偶热提起说topo克隆很困难，我让她用
promega的，解决。
由于克隆做不出来，我们实验室在那半年反复定kit，短期内invitrogen的销量可能反
而增加。对顾客反映问题如此消极对待，真的很差劲,公司可能反而会为销量增加沾沾
自喜。
版上如果有遇到topo不好用的，也写信反映一下。如果多人反映还不解决，那就在哪里
曝光一下它。
提供试剂的公司产品不稳定是非常严重的问题，浪费一大批人的时间。 |
|
r****r
发帖数: 379 | 23 规模大了以后，Invitrogen雇佣了一大批GE的完全不懂生物技术的人管理公司，内部的
技术管理出了许多问题，有些产品的QC也出问题，他们内部用起来抱怨也很多。
其实大家反应的也基本属实，我认识invitrogen market部门的，他们叫卖的东西不是
cloning products，大部分算是二线品牌吧，谈不上最好，RT－PCR system还不错，我
一直用，但现在也不用了，贵，不划算。 |
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r**h
发帖数: 6 | 24 Thanks. Good to know all about your experiences and thoughts.
I guess that the sequences with apparently essential roles could be quite
conserved between 129 and B6, say, protein coding sequences, and perhaps the
promoter region you mentioned (certainly I need to carefully check with
this claim). This could be one of the reasons why you were able to go with
B6, a fortunate move. So, maybe I should take my chance to grab a B6 clone,
because the region I am eyeing for targeting is pretty close to |
|
b***g
发帖数: 516 | 25 哪些地方可以购买BAC clones阿？Open biosystems?
Thanks! |
|
a****d
发帖数: 1919 | 26 从起始密码子开始，序列是这样的：
ATGCCGGCCGGCCGCGCCGCGCGC
gene本身65％GC含量,1.3Kb
打算从ES cell的cDNA pool中clone到lenti over expression vector，ES cell的cDNA
pool是从mRNA反转录得到的。目前试过TAKARA的LA taq，GC buffer，with or
without DMSO(5%),做了gradient Tm， 从53－58度，结果任何一个条件都不能PCR出片
断。现在想是不是因为起始序列太特别了，有没有特别的办法把这个ORF扩增出来？ |
|
s******r
发帖数: 2876 | 27 你先看看这个基因有没有的卖，用纯的plasmid作template容易一点。
在你可以考虑在atg之前寻找合适的primer，先得到PCR product
然后再PCR ORF。
roche有一个high gc的PCR kit，用过还行。
从起始密码子开始，序列是这样的：
ATGCCGGCCGGCCGCGCCGCGCGC
gene本身65％GC含量,1.3Kb
打算从ES cell的cDNA pool中clone到lenti over expression vector，ES cell的cDNA
pool是从mRNA反转录得到的。目前试过TAKARA的LA taq，GC buffer，with or
without DMSO(5%),做了gradient Tm， 从53－58度，结果任何一个条件都不能PCR出片
断。现在想是不是因为起始序列太特别了，有没有特别的办法把这个ORF扩增出来？ |
|
g*********5
发帖数: 2533 | 28 you could clone part of this gene into plasmid first, then insert mutation. |
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m****n
发帖数: 1066 | 29 正在clone一段大约2000bp的 3’UTR.
1. RT-PCR, 用过superscriptIII and thermoscript, 不行。靠近polyA的可以PCR出来
。这个2000bp的不行。
2. 用human genomics DNA, 自己从细胞里用DNAeasy提的。至少三四条带在1500-
2000bp的位置。
这个序列二级结构比较多。
高手指点。不行就找genescript合成了。 |
|
b******e
发帖数: 61 | 30 多谢楼上的回复。
因为我的目的是想把4kb片断弄到pLenti7.3里边去，没有合适的酶切位点，只好用TA
cloning. 我也试过不用kit里边带的topo, 自己用T4 Ligase做TA连接，也失败了。 |
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r*****m
发帖数: 231 | 31 把需要的位点设计在PCR引物的两端，PCR后用酶切然后连上vector。你的Lenti vector
应该至少有7，8kb吧，连个4kb的一般不会出问题。一般人做PCR cloning都是这么做的
，做不出来的才用TOPO。但你是TOPO都做不出来，就不知道为什么了。。。检查下你的
Taq是不是有问题吧。。。 |
|
X***n
发帖数: 366 | 32 genome.ucsc.edu
open the BAC track. Go to gene ApoE. Select the BAC clone. You can order
from Invitrogen.
and
non- |
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t******r
发帖数: 8600 | 33 要克隆一基因locus,30-50 kb. Genbank没有sequence.大量repetitive segments, PCR
基本不work.好不容易PCR出一4kb片段，TA-cloning总被重组掉80-90%。试用过不同菌
种，30C培养等等。求高人指点。 |
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t******r
发帖数: 8600 | 34 Fosmid & BAC libray都做了三个了。能筛出其他基因，但从来没找到这clone的影子。
没有
genomic probe, 只有cDNA probe.
PCR是分段进行的。 |
|
p****p
发帖数: 3360 | 35 Hi, there,
I'm a fruit fly people and want to order a human cDNA clone from either the
NIH_MGC_43 or NIH_MGC_114 library.
Where's the best place to order it?
Any information is appreciated. Thanks a lot in advance. |
|
M******t
发帖数: 555 | 36 129 mice strain 的 bac clone 大家在哪里order的？ |
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m*********r
发帖数: 136 | 37 I try to clone a gene by cutting it from one vector to another. However, the
restriction enzyme site will also cut the polyA from vector A, while vector
B already has the SV40 polyA. Is that ok to have two polyA sequence in the
vector?
scheme is like this
MCS+gene+polyA---junk sequence--polyA--
Best, |
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v***a
发帖数: 1242 | 38 要overexpress一个蛋白（full length），在NCBI查它的nucleotide sequence来设计
cloning primer，发现有两个看起来都可信的reference sequence，blast了一下，发
现这两个sequence在CDS区有一部分是相同并连贯的，其它就都不同了。其中sequence
1只是cds，并且是partial cds，没有stop codon；sequence 2是complete cds，包括
非CDS区，好像是由NIH MGC (Mammalian Gene Collection) Project来的，但由
sequence 1衍生出的AA序列却比sequence 2衍生的AA序列还要多50多个AA。
我选择了全长的sequence 2设计primer，一共设计了3对，PCR后都未能得到预期bp的
band（虽然有的PCR出来是single band），我每对primer都挑了最接近预期长度的PCR
product去sequence，但都不是我想要的gene。。。难道是sequence 2本身就有问题么
？（我是挑的非CDS区 |
|
c******r
发帖数: 3778 | 39 你是说2kb？如果你想表达这个蛋白，很简单啊。设计对primer，提点RNA直接做RT-PCR
好了。这样得到的就是coding sequence，没有intron，表达比较容易吧？
从genomic DNA上clone除了exon还有intron，有时候不一定在细胞里面能够正确表达。 |
|
n********k
发帖数: 2818 | 40 pretty easily to deal with that, and if you get the luck, u might even
easily publish a small paper about it along
the cloning... |
|
v***a
发帖数: 1242 | 41 真是非常感谢你！你给的答案就是我想问的！
你所说的前两步我已经做了，所以已知此蛋白表达，并且sequence 2很可能不适用于我
想要得到的组织中的gene。
后来感谢clonebar同学，他帮我查到了一个序列，跟我查到的sequence 1是一致的，并
且有一部分的5'及3' UTR sequence，我刚设计了新的primer，下周到就可以进行实验
了。
我也几乎翻遍了跟我这个gene有关的paper，没有见到有clone full-length的，并且
sequence 2貌似是大家引用最多的（比如设计此gene的siRNA等）。
谢谢你说的5'和3' RACE，这个我没想到，如果sequence 1也不行，我就会采取这个办
法。
非常感谢！！！
2
隆。
region
2
800 |
|
o********r
发帖数: 775 | 42 十年前俺做过这事（17Kb的plasmid），而且过程和你说的很象，最后筛了5个月（每天
就是早上抽质粒，下午PCR/跑胶，然后ligation/transformation，回家前pick colony
/incubation O/N），前后尝试了无数种ligase/competent cell，几乎市面上相关产品
都试了个遍，终于找到一个要的clone。其实17Kb对于某些质粒而言已经差不多是上限
，不稳定，体内很容易发生重组，如果可能尝试不同的vector，或者不同的competent
cell/transformation protocol。另外你或许可以试试Ca++ based
transformation。 |
|
D******9
发帖数: 2665 | 43 我有个研究的基因有很多DIFFERENT ISOFORMS BECAUSE OF ALTERNATIVE SPLICING.我
想把所有的IOFORMS都CLONE了，现在NCBI数据库了不全， 除了筛选cDNAlibrary的方法
， 还有别的方法可以拿到所有的序列吗？ 我只知道基因中间的一段。谢谢 |
|
b******r
发帖数: 111 | 44 I am cloning 3.4kb human genome DNA into a 8.4kb vector.Isolate some
plasmids. Recut,then 3.4kb bands
appear. But sequencing shows it is not 3.4kb human DNA,it is a unknown
fragment. Strangely,the size is same
as 3.4kb! What happen? Any idea to eliminate recombination ? The competent
cells used are Invitrogen ElectroMAX™ DH10B™ T1 Phage-
Resistant Competent Cells Cat. No. 12033-015. |
|
