c*****t
发帖数: 1879 | 1 Two methods that I could think of as possibility:
1. How about F' cells? The F' circle can be treated as a plasmid...
They can recombine with bacterial chromosome or mate w/ other
individual bacteria.
2. Obtain crude cell extract and try to find fractions that still has
DNA polymerase activity. Then use Sanger's method (correct spelling?)
to sequence DNA using a random/degenerate primer (extend the primer
until only one sequence is obtained...)..
Please post the correct answer... | g******k
发帖数: 188 | 2
I don't know the correct answer either, that is why I am asking,
one of my committee member asked me about this
I think there are some tricky part, he mentioned about using lacZ opern
this is realy a tough question for those whose major in Molecular Biology.May
be only some guys from Harvard or MIT etc. could answer it,or it is very bad
question.
the question is"how would you clone a gene in 1950's, when the endonuclease
has not been dicovered yet?"
【在 c*****t 的大作中提到】
: Two methods that I could think of as possibility:
: 1. How about F' cells? The F' circle can be treated as a plasmid...
: They can recombine with bacterial chromosome or mate w/ other
: individual bacteria.
: 2. Obtain crude cell extract and try to find fractions that still has
: DNA polymerase activity. Then use Sanger's method (correct spelling?)
: to sequence DNA using a random/degenerate primer (extend the primer
: until only one sequence is obtained...)..
: Please post the correct answer...
| s**********i
发帖数: 711 | 3
3 people won 1978 Nobel price for discovering endonuclease.
any paper on molecular cloning before early 70s should have no
endonuclease mentioned...
【在 g******k 的大作中提到】
:
: I don't know the correct answer either, that is why I am asking,
: one of my committee member asked me about this
: I think there are some tricky part, he mentioned about using lacZ opern
: this is realy a tough question for those whose major in Molecular Biology.May
: be only some guys from Harvard or MIT etc. could answer it,or it is very bad
: question.
: the question is"how would you clone a gene in 1950's, when the endonuclease
: has not been dicovered yet?"
| k***r
发帖数: 100 | 4 One possible way is using Bac. Phage. Since it can random pick up Bac. genome. So
you get a lirary of phage with random insert, use it to do complement test,
see which one will recover the lost function. But the problem is once you get
the phage that has right insert DNA, how do you get it out? Or you have to do
Sanger's seq. through the phage genome (which is impossible at 50s'):(
【在 g******k 的大作中提到】
:
: I don't know the correct answer either, that is why I am asking,
: one of my committee member asked me about this
: I think there are some tricky part, he mentioned about using lacZ opern
: this is realy a tough question for those whose major in Molecular Biology.May
: be only some guys from Harvard or MIT etc. could answer it,or it is very bad
: question.
: the question is"how would you clone a gene in 1950's, when the endonuclease
: has not been dicovered yet?"
| c*****g
发帖数: 1856 | 5 tho u can use mechnic method to break the DNA and then use the laz to pick
the gene fragment. maybe this can be done . still have many problem in d
detail.
【在 g******k 的大作中提到】
:
: I don't know the correct answer either, that is why I am asking,
: one of my committee member asked me about this
: I think there are some tricky part, he mentioned about using lacZ opern
: this is realy a tough question for those whose major in Molecular Biology.May
: be only some guys from Harvard or MIT etc. could answer it,or it is very bad
: question.
: the question is"how would you clone a gene in 1950's, when the endonuclease
: has not been dicovered yet?"
| c*****t
发帖数: 1879 | 6 I asked a microbiol professor. He gave me a possibility:
if you want to clone an antibiotic resistance gene, it can be
done by increasing the dosage of antibiotics, for instance, cam,
in order to clone (ie having more than 1 copy and on a plasmid).
Only cells w/ plasmids that can produce high copy can survive.
coco
【在 g******k 的大作中提到】
:
: I don't know the correct answer either, that is why I am asking,
: one of my committee member asked me about this
: I think there are some tricky part, he mentioned about using lacZ opern
: this is realy a tough question for those whose major in Molecular Biology.May
: be only some guys from Harvard or MIT etc. could answer it,or it is very bad
: question.
: the question is"how would you clone a gene in 1950's, when the endonuclease
: has not been dicovered yet?"
|
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