Abstract
| Originalsprog | Engelsk |
|---|---|
| Tidsskrift | Mitochondrial D N A |
| Vol/bind | 11 |
| Udgave nummer | 3 & 4 |
| Sider (fra-til) | 183-192 |
| ISSN | 1940-1736 |
| DOI | |
| Status | Udgivet - 2000 |
Bibliografisk note
Keywords: Sulfolobus; genome mapping; paired-PCR; BAC clonesAdgang til dokumentet
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I: Mitochondrial D N A, Bind 11, Nr. 3 & 4, 2000, s. 183-192.
Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › peer review
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TY - JOUR
T1 - A Bac Library and Paired-PCR Approach to Mapping and Completing the Genome Sequence of Sulfolobus Solfataricus P2
AU - She, Qunxin
AU - Confalonieri, F.
AU - Zivanovic, Y.
AU - Medina, N.
AU - Billault, A.
AU - Awayez, Mariana J.
AU - Thingoc, Hoa Phan
AU - Thi Pham, Boi-Tien
AU - van der Oost, J.
AU - Duguet, M.
AU - Garrett, Roger A.
N1 - Keywords: Sulfolobus; genome mapping; paired-PCR; BAC clones
PY - 2000
Y1 - 2000
N2 - The original strategy used in the Sulfolobus solfatnricus genome project was to sequence non overlapping, or minimally overlapping, cosmid or lambda inserts without constructing a physical map. However, after only about two thirds of the genome sequence was completed, this approach became counter-productive because there was a high sequence bias in the cosmid and lambda libraries. Therefore, a new approach was devised for linking the sequenced regions which may be generally applicable. BAC libraries were constructed and terminal sequences of the clones were determined and used for both end mapping and PCR screening. The PCR approaches included a novel chromosome walking method termed “paired-PCR”. 21 gaps were filled by BAC end sequence analyses and 6 gaps were filled by PCR including three large ones by paired-PCR. The complete map revealed that 0.9 Mb remained to be sequenced and 34 BAC clones were selected for walking over small gaps and preparing template libraries for larger ones. It is concluded that an optimal strategy for sequencing microorganism genomes involves construction of a high-resolution physical map by BAC end analyses, PCR screening and paired-PCR chromosome walking after about half the genome sequence has been accumulated.
AB - The original strategy used in the Sulfolobus solfatnricus genome project was to sequence non overlapping, or minimally overlapping, cosmid or lambda inserts without constructing a physical map. However, after only about two thirds of the genome sequence was completed, this approach became counter-productive because there was a high sequence bias in the cosmid and lambda libraries. Therefore, a new approach was devised for linking the sequenced regions which may be generally applicable. BAC libraries were constructed and terminal sequences of the clones were determined and used for both end mapping and PCR screening. The PCR approaches included a novel chromosome walking method termed “paired-PCR”. 21 gaps were filled by BAC end sequence analyses and 6 gaps were filled by PCR including three large ones by paired-PCR. The complete map revealed that 0.9 Mb remained to be sequenced and 34 BAC clones were selected for walking over small gaps and preparing template libraries for larger ones. It is concluded that an optimal strategy for sequencing microorganism genomes involves construction of a high-resolution physical map by BAC end analyses, PCR screening and paired-PCR chromosome walking after about half the genome sequence has been accumulated.
U2 - 10.3109/10425170009033231
DO - 10.3109/10425170009033231
M3 - Journal article
SN - 1940-1736
VL - 11
SP - 183
EP - 192
JO - Mitochondrial D N A
JF - Mitochondrial D N A
IS - 3 & 4
ER -