Re: 大家看看吧
※ 引述《weisont (我的腰好酸)》之銘言：
: 這應該是兩回事吧
: PCR是PCR
: CLONE是CLONE
: 兩者並不是有相同功能而可以互相取代的
: PCR是將目標基因放大
: CLONE的目的一部分是保存目標基因
: 一部分則是做INVIVO的實驗
: 如果不做CLONE
: 那要怎麼來表現這段基因勒??
喔 原來我剛剛誤會了~~
[因為我太累 懶的看英文...||]
原來呢..作clone的目的 是想得到這段基因表現出來的rRNA
然後比較rRNA的特殊二級結構
就可以確定PCR的過程中有沒有出狀況..
但是因為現在PCR的技術越來越好
所以已經不用透過這個步驟來confirm了...
嗯..應該是吧..[我還是沒有看下面那篇文章...||]
所以 有問題再提出來吧XD
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以下是資料的原文..
In summary, there appears to be no reason for more than
the usual concern, because of Taq polymerase fidelity of
replication, about sequence accuracy in this method. Several
positions that had previously been scored as unknown or
ambiguous are now determinable by taking advantage of
bidirectional gene sequencing and the absence of artifacts
due to rRNA secondary structure and posttranscriptionally
modified nucleotides. As always, a cautious approach should
be taken by checking any sequence anomalies and confirming
known secondary structural constraints on sequences.
Although we have spent considerable time optimizing and
testing the direct sequencing of PCR-amplified rDNA, we
highly recommend cloning the fragments if a near-perfect
sequence is desired.
........
The amplification by PCR of a taxonomically diverse collection
of eubacterial 16S rDNA genes is possible with a small number
of primers. These products can readily be cloned for sequencing
or they can be sequenced directly.
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