Sequence and locational analysis of qumA-qumB gene cluster specific to Xanthomonas arboricola

• Main Authors: Sue-Lin Chou, 周淑玲
• Other Authors: Yung-An Li
• Format: Others
• Language: en_US
• Published: 2000
• Online Access: http://ndltd.ncl.edu.tw/handle/87709126269943555720

碩士 === 輔仁大學 === 生物學系 === 88 === 英文摘要 The five pathovars （X. arboricola pv. carotae、X. arboricola pv. celebensis、X. arboricola pv. corylina、X. arboricola pv. juglandis以及X. arboricola pv. pruni）of Xanthomonas arboricola that metabolize quinate were discovered. A gene involved in quinate metabolism had been cloned from X. arboricola pv. juglandis strain XJC5. The gene, qumA, located on 22.1 kb SQ2 clone from X. arboricola pv. juglandis strain XJC5, conferred quinate metabolic activity to X. arboricola pv. celebensis XC143 （SQ（-）strains）. In this report, another gene-qumB, nearing the 3’-end of qumA, had the same transcriptional direction with qumA and showed specific to the five X. arboricola pathovars in Xanthomonas. Therefore, the nucleotide sequence of qumA and qumB were used to design two kinds of specific primer pairs（SQ-F2+SQ-R2 and Porin-F2+Porin-R） to distinguish X. arboricola from the other Xanthomonas genomic DNA. On the other hand, we also used two primer pairs （Q-F’+Porin-R’） to prove that qumA and qumB got together （qumA-qumB gene cluster） in the five X. arboricola pathovars by PCR （polymerase chain reaction）. On the both sides of qumA-qumB, we found mem （membrane protein gene）、unknown ORF （uncertainly gene） and pep （peptidase gene）were together clustered in the order mem-unknown-qumA-qumB-pep and they had the same transcriptional direction in X. arboricola pv. juglandis strain XJC5. The genes homology to mem、unknown ORF and pep from X. arboricola pv. juglandis strain XJC5 existed not only in X. arboricola by Southern hybridization. In other words, almost all of the Xanthomonas had the three genes homology to mem、unknown ORF and pep in X. arboricola pv. juglandis strain XJC5. Therefore, we designed the other two kinds of primer pairs （X-F’+Q-R’ and Porin-F’+Pep-R’） to ensure that qumA-qumB in the five Xanthomonas pathovars were all between unknown and pep. This result indicated qumA-qumB gene cluster was so specific to X. arboricola that it is possible came from foreign biology into Xanthomonas. According to the five probes, the internal fragments of mem、unknown ORF 、qumA、qumB and pep, we could division 33 pathovars of Xanthomonas into six classes：class I contained X. hortorum pv. pelargonii strain XP15(T) and X. punicae strain XP177(T) which could not hybrid to all of the five probes；class II contained X. campestris pv. mangiferaeindicae strain XCM28-2 which could only hybrid to unknown probe；class III contained X. arrhenatheri strain XA(T)、X. poae strain XP(T) and X. phlei strain XP(T) （belong to X. campestris DNA homology group 3） which could only hybrid to mem and pep probes；class IV contained X. eucalypti strain XE104(T) which could only hybrid to mem and unknown probes；the other which could only hybrid to mem、unknown and pep probes were called class V；class VI was the X. arboricola which could hybrid to all of the five probes. Using the five probes, we could distinguish XCM28-2 and XE104(T) from the other 31 pathovars of Xanthomonas genomic DNA by Southern hybridization. If we collect enough probes in the future, maybe we can use this method to distinguish all of the Xanthomonas pathovars. Because of qumA-qumB was specific to the five X. arboricola pathovars and was not found in the other Xanthomonas species, we suspected that the qumA-qumB gene cluster came from other plant pathogens. To understood whether other plant pathogens also had the ability to metabolize quinate. Our laboratory tested some plant pathogens of Erwinia and Pseudomonas. We found that E. ananas、E. herbicola、E. herbicola pv. millettiae、E. rhapontici以及E. cypripedii （belong to Erwinia cluster I and cluster II） have the same color change in SQY medium （dark green） and P. syringae have the different color change in SQY medium （yellowwish）. Moreover, the qumA probe could hybrid to all the test Erwinia belong to Erwinia cluster I、cluster II strains and two pathovars of P. syringae and qumB probe could only hybrid to E. rhapontici strain ER1 （belong to Erwinia cluster II） alone so far tested. So that qumA or qumB at least existed between the three kinds of plant pathogens, and might be took place horizontal transfer between them.