avatar
about sequencing pCMV# Biology - 生物学
h*g
1
I bought a pCMVsport6 vector with one gene inserted. I want to confirm the
sequence of the gene. Using T7 primers, I can get part of the sequence. But
using sp6, M13 forward and reverse primers, I cannot get any sequence. Why?
Can anyone help me about this?
Thanks.
avatar
l*a
2
不能用cmv protomoter 来sequence?

But
?

【在 h*********g 的大作中提到】
: I bought a pCMVsport6 vector with one gene inserted. I want to confirm the
: sequence of the gene. Using T7 primers, I can get part of the sequence. But
: using sp6, M13 forward and reverse primers, I cannot get any sequence. Why?
: Can anyone help me about this?
: Thanks.

avatar
l*y
3
cmv的sequence google一下就有了,很多测序公司都有cmv的引物
实在不行用你gene的sequence去搞几个primer测几段overlap的自己拼

But
?

【在 h*********g 的大作中提到】
: I bought a pCMVsport6 vector with one gene inserted. I want to confirm the
: sequence of the gene. Using T7 primers, I can get part of the sequence. But
: using sp6, M13 forward and reverse primers, I cannot get any sequence. Why?
: Can anyone help me about this?
: Thanks.

avatar
h*g
4
I have not tried cmv promoter. But sp6 or M13 reverse is more closer to the
3' end of my gene. Any suggestions? Thanks.

【在 l******a 的大作中提到】
: 不能用cmv protomoter 来sequence?
:
: But
: ?

avatar
h*g
5
Next I will use restriction enzyme to cut, and use T7 and SP6 to make probes
. So I want to know the correct sequence. Why sp6, M13forward and reverse
not working? Thanks.

But
?

【在 h*********g 的大作中提到】
: I bought a pCMVsport6 vector with one gene inserted. I want to confirm the
: sequence of the gene. Using T7 primers, I can get part of the sequence. But
: using sp6, M13 forward and reverse primers, I cannot get any sequence. Why?
: Can anyone help me about this?
: Thanks.

avatar
T*e
6
不知道有个小细节楼主注意过没有,有的时候通用引物的序列可能是反的
不过你说完全没有测出来,是引物有问题吗?
如果你有那个插入的gene序列的话,我建议你自己每隔500bp设计一个引物,把这个基
因测通
avatar
f*f
7
买的载体总会有序列,公司的通用引物也该有序列吧。比对一下看看具体在哪有没问题。
-------------------
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是指sequence of insertion还是完全没信号,假如引物跟载体能匹配的话估计就是测
序的问题了,想测哪段自己合成引物也成。
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