What is the distinction between F' plasmid and R plasmid?

What is the distinction between F' plasmid and R plasmid?

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Is there a difference between an F' plasmid that has taken up a chromosomal gene that conveys antibiotic resistance, and an R plasmid? Is a bacterium containing an R plasmid and yet lacking an F+ plasmid capable of horizontal gene transfer by conjugation?

If I understand the nomenclature correctly, an R plasmid is just any plasmid containing an antibiotic (R)esistance gene (eg. Amp, Kan, Cm, etc.). It's a bit of an outdated name from when people didn't know how exactly the plasmids conferred such resistance.

An F-plasmid is any plasmid that contains the genes necessary for (F)ertility, eg:horizontal gene transfer via the tra operon.

The two do not have to appear together -- eg: Many F plamids will contain resistance genes (along with other random chunks from the genome), but resistance doesn't always have to be part of the payload. Likewise, it is common in labs to build pure resistance vectors that lack any horizontal transfer capability in order to select for particular clones.

Recognize the characteristics of, and thus the functions, of plasmids

  • The gene to be replicated is inserted into copies of a plasmid containing genes that make cells resistant to particular antibiotics, and into a multiple cloning site (MCS, or polylinker), allowing the easy insertion of DNA fragments.
  • A major use of plasmids is to make large amounts of proteins. Bacterium can be induced to produce large amounts of proteins from the inserted gene. This is a cheap and easy way of mass-producing a gene or the protein it then codes for for example, insulin or even antibiotics.
  • It is possible for plasmids of different types to coexist in a single cell. Several different plasmids have been found in E. coli. However, related plasmids are often incompatible, in the sense that only one of them survives in the cell line, due to the regulation of vital plasmid functions.

Structure and Composition

Plasmids are small molecules of double stranded, helical, non-chromosomal DNA. In most plasmids the two ends of the double-stranded DNA molecule that make up plasmids covalently bond together forming a physical circle. Some plasmids, however, have linear DNA. Plasmids replicate independently of the host chromosome, but some plasmids, called episomes, are able to insert or integrate into the host cell&rsquos chromosome where their replication is then regulated by the chromosome.

Although some plasmids can be transmitted from one bacterium to another by transformation and by generalized transduction, the most common mechanism of plasmid transfer is conjugation. Plasmids that can be transmitted by cell-to-cell contact are called conjugative plasmids. They contain genes coding for proteins involved in both DNA transfer and and the formation of mating pairs.

General Types of Plasmids

Conjugative and Non-Conjugative

There are many ways to classify plasmids from general to specific. One way is by grouping them as either conjugative or non-conjugative. Bacteria reproduce by sexual conjugation, which is the transfer of genetic material from one bacterial cell to another, either through direct contact or a bridge between the two cells. Some plasmids contain genes called transfer genes that facilitate the beginning of conjugation. Non-conjugative plasmids cannot start the conjugation process, and they can only be transferred through sexual conjugation with the help of conjugative plasmids.


Another plasmid classification is by incompatibility group. In a bacterium, different plasmids can only co-occur if they are compatible with each other. An incompatible plasmid will be expelled from the bacterial cell. Plasmids are incompatible if they have the same reproduction strategy in the cell this allows the plasmids to inhabit a certain territory within it without other plasmids interfering.

Conjugation and Compatibility

Conjugation is a process by which bacterial plasmid can be transferred from one cell to another cell. Conjugative plasmids involve in the process of sexual conjugation. On the other hand, non-conjugative plasmids are also present in bacterial cells. The non-conjugative plasmids lack a type of gene called tra gene which is involved in the transfer of plasmids by conjugation and present in conjugative plasmids. A bacterial cell can have several types of plasmids among which different types of conjugative plasmids are also present. In this case, the transfer process required the compatibility of the plasmids present in a bacterial cell. Two incompatible plasmids will not be present inside a cell one or the other will be lost. Hence, the compatibility of the plasmids plays an important role in the selection of cloning vectors.

Fertility Plasmids

Like many other plasmids, fertility plasmids (F plasmid) have a circular structure and measures about 100 kb.

Some of the main parts of the F plasmid include:

  • Transposable element (IS2, 1S3, and Tn1000)
  • Replication sites (RepFIA, RepFIB, and RepFIC)
  • Origin of conjugative transfer (oirT)
  • Replication origin regions

F plasmid plays an important role in reproduction given that they contain genes that code for the production of sex pilus as well as enzymes required for conjugation. F plasmid also contains genes that are involved in their own transfer. Therefore, during conjugation, they enhance their own transfer from one cell to another.

Whereas the cells that process the F plasmids are referred to as donors, those that lack this factor are the recipients. On the other hand, the plasmids that enhance the ability of the host cell to behave like a donor are known as the transfer factor.

During conjugation, the donor cell (bacteria) with sex pili (1-3 sex pili) binds to a specific protein on the outer membrane of the recipient thus initiating the mating process.

Following the initial binding, the pili retract thus allowing the two cells to bind together. This is then followed by the transfer of DNA from the donor to the recipient and consequently the transfer of the F plasmid. As a result, the recipient acquires the F factor and gains the ability to produce sex pilus involved in conjugation.

* During conjugation, only DNA is passed from the donor to the recipient. Therefore, cytoplasm and other cell material are not transferred.

* Sexual pili are tiny rod-like structures that allow the F-positive (cells that have the F factor) bacterial cells to attach to the F-negative (cells lacking the pili) female to promote conjugative transfer.

What is the distinction between F' plasmid and R plasmid? - Biology

ຄວາມແຕກຕ່າງທີ່ ສຳ ຄັນລະຫວ່າງ F plasmid ແລະ R plasmid ແມ່ນວ່າ F plasmid ແມ່ນ DNA extrachromosomal ເຊິ່ງມີລະຫັດພັນທຸ ກຳ ສຳ ລັບປັດໄຈການຈະເລີນພັນ. ໃນຂະນະດຽວກັນ, R plasmid ແມ່ນ DNA extrachromosomal ເຊິ່ງມີລະຫັດພັນທຸ ກຳ ສຳ ລັບການຕໍ່ຕ້ານກັບຢາຕ້ານເຊື້ອ.

plasmid ແມ່ນ DNA ຂະ ໜາດ ນ້ອຍວົງສອງວົງທີ່ມີຢູ່ໃນແບັກທີເຣຍ. ພວກມັນແມ່ນ DNA extrachromosomal ແລະມີຄວາມສາມາດໃນການທົດແທນຕົວເອງ. ພວກມັນປະຕິບັດພັນທຸ ກຳ ທີ່ ຈຳ ເປັນ ສຳ ລັບການ ຈຳ ລອງແລະຮັກສາຕົວເອງ. ນອກຈາກບັນຈຸພັນທຸ ກຳ ທີ່ ຈຳ ເປັນ ສຳ ລັບການ ຈຳ ລອງຕົວເອງ, plasmids ຍັງມີອີກຫຼາຍໆຊະນິດທີ່ ຈຳ ເປັນ ສຳ ລັບການລະຫັດຄຸນລັກສະນະພິເສດເຊັ່ນການຕ້ານທານຕ້ານເຊື້ອ, ການເຊື່ອມໂຊມຂອງ macromolecules, ຄວາມທົນທານຂອງໂລຫະ ໜັກ, ການຜະລິດ bacteriocins, ການໂອນ ກຳ ມະພັນແລະອື່ນໆເຊິ່ງເປັນ ມີປະໂຫຍດຕໍ່ເຊື້ອແບັກທີເຣັຍ.

ຍິ່ງໄປກວ່ານັ້ນ, ມີ plasmids ທີ່ຫລາກຫລາຍ. R plasmids ແລະ F plasmids ແມ່ນສອງຊະນິດໃນນັ້ນ. F plasmid ແມ່ນ plasmid ທີ່ມີຄວາມສາມາດໃນການຜະສົມຜະສານແລະການຜະລິດ pili ທາງເພດ. R plasmid ແມ່ນ plasmid ຕ້ານທານທີ່ມີຄວາມສາມາດໃນການຕໍ່ຕ້ານກັບຢາຕ້ານເຊື້ອແລະຕົວຍັບຍັ້ງການຈະເລີນເຕີບໂຕຂອງເຊື້ອແບັກທີເລຍບາງຊະນິດ.


1. ພາບລວມແລະຄວາມແຕກຕ່າງທີ່ ສຳ ຄັນ 2. What is F Plasmid 3. What is R Plasmid 4. ຄວາມຄ້າຍຄືກັນລະຫວ່າງ F Plasmid ແລະ R Plasmid 5. ການປຽບທຽບຂ້າງຄຽງ - F Plasmid vs R Plasmid ໃນແບບຟອມ Tabular 6. Summary

F Plasmid ແມ່ນຫຍັງ?

ສາຍພັນແບັກທີເລຍບາງຊະນິດມີ plasmids F ນອກ ເໜືອ ໄປຈາກໂຄໂມໂຊມຂອງພວກມັນ. ສາຍພັນເຫຼົ່ານີ້ເອີ້ນວ່າສາຍພັນ F +. ພວກມັນເຮັດ ໜ້າ ທີ່ເປັນຈຸລັງບໍລິຈາກຫລືຊາຍໃນການຕ້ານເຊື້ອແບັກທີເຣຍເຊິ່ງເປັນກົນໄກການສືບພັນທາງເພດທີ່ສະແດງໂດຍເຊື້ອແບັກທີເຣັຍເຊິ່ງ ອຳ ນວຍຄວາມສະດວກໃຫ້ແກ່ການໂອນຍ້າຍແນວນອນໃນລະຫວ່າງແບັກທີເຣຍ. F plasmids ສາມາດ ຈຳ ລອງແບບອິດສະຫຼະແລະບັນຈຸປັດໄຈການຈະເລີນພັນທີ່ມີຊື່ວ່າ gen tra. ສະນັ້ນ, DNA extrachromosomal (plasmids) ເຫຼົ່ານີ້ມີຊື່ວ່າ F plasmids ເນື່ອງຈາກປັດໄຈ F ຫຼືປັດໄຈການຈະເລີນພັນ. ກຳ ມະພັນລະຫັດປັດໄຈທາງພັນທຸ ກຳ ແມ່ນມີຄວາມ ຈຳ ເປັນ ສຳ ລັບການໂອນຖ່າຍຫລືການເຊື່ອມຕົວ.

ສາຍພັນແບັກທີເຣຍທີ່ໄດ້ຮັບ plasmids F ຈາກສາຍພັນ F + ແມ່ນຮູ້ກັນວ່າສາຍພັນ F ຫຼືສາຍພັນຂອງຜູ້ຮັບຫລືຜູ້ຍິງ. ສາຍພັນ F + ສາມາດບໍລິຈາກເອກະສານພັນທຸ ກຳ ຂອງເຂົາເຈົ້າຫຼື DNA extrachromosomal ໃຫ້ກັບເຊື້ອແບັກທີເລຍອື່ນ.

ການຕ້ານເຊື້ອແບັກທີເຣຍເລີ່ມຕົ້ນຈາກການຜະລິດ pili ທາງເພດ ສຳ ພັນໂດຍສາຍພັນ F + ເພື່ອຕິດຕໍ່ກັບເຊື້ອແບັກທີເຣັຍ F. ເພດທົດລອງຊ່ວຍ ອຳ ນວຍຄວາມສະດວກໃຫ້ເຊນການສື່ສານແລະການຕິດຕໍ່ຫ້ອງໂດຍການສ້າງທໍ່ເຊື່ອມ. ການສ້າງຕັ້ງນີ້ແມ່ນຄວບຄຸມໂດຍເຊື້ອສາຍປັດໄຈການຈະເລີນພັນທີ່ເກີດມາຈາກສາຍພັນ F +. F + ຈຳ ລອງ F plasmid ຂອງມັນແລະເຮັດ ສຳ ເນົາມັນເພື່ອໂອນເຂົ້າ F-strain. ສຳ ເນົາ F plasmid ທີ່ໂອນໄປ F-strain ຜ່ານທໍ່ເຊື່ອມ. ເມື່ອໃດທີ່ມັນໂອນ, ທໍ່ການເຊື່ອມຕົວເຂົ້າກັນ. ສາຍຜູ້ຮັບຈະກາຍເປັນ F +. ໃນລະຫວ່າງການປະສົມເຊື້ອແບັກທີເລຍ, ພຽງແຕ່ F plasmid ຖືກໂອນຈາກສາຍພັນ F + ໄປ F-strain ໂດຍບໍ່ຕ້ອງໂອນໂຄໂມໂຊມແບັກທີເຣຍ.

R Plasmid ແມ່ນຫຍັງ?

R plasmid ຫຼືພູມຕ້ານທານ plasmid ແມ່ນ DNA ທີ່ມີໂຄໂມໂຊມພິເສດຂອງເຊື້ອແບັກທີເຣັຍທີ່ມີລະຫັດພັນທຸ ກຳ ສຳ ລັບການຕໍ່ຕ້ານກັບຢາຕ້ານເຊື້ອ. ເພາະສະນັ້ນ, plasmids R ທີ່ມີເຊື້ອແບັກທີເຣັຍສະແດງໃຫ້ເຫັນການຕໍ່ຕ້ານກັບຢາຕ້ານເຊື້ອ. ນັກວິທະຍາສາດຍີ່ປຸ່ນໄດ້ພົບເຫັນ plasmids R ຖືກສະແດງໃຫ້ເຫັນໃນເຊື້ອແບັກທີເຣັຍ Shigella ກ່ອນ. R plasmids ແມ່ນເປັນທີ່ຮູ້ຈັກເປັນປັດໃຈ R ກ່ອນທີ່ຈະເຂົ້າໃຈເຖິງລັກສະນະຂອງ plasmids. ໂດຍທົ່ວໄປແລ້ວ, plasmids R ມີຫລາຍພັນທຸ ກຳ ທີ່ຕ້ານທານກັບຢາຕ້ານເຊື້ອ. ເວົ້າອີກຢ່າງ ໜຶ່ງ, ລະຫັດປັດໄຈ R ດຽວ ສຳ ລັບເຊື້ອພັນທີ່ຕ້ານທານກັບຢາຕ້ານເຊື້ອຫຼາຍກວ່າ, ບາງຄັ້ງມີເຖິງ 8 ໂຕຕ້ານເຊື້ອແຕກຕ່າງກັນ.

ພູມຕ້ານທານຂອງຢາຕ້ານເຊື້ອຫຼື plasmids R ສາມາດແຜ່ລາມຈາກເຊື້ອແບັກທີເຣຍ ໜຶ່ງ ໄປສູ່ອີກເຊື້ອ ໜຶ່ງ ແລະແຜ່ລາມຜ່ານເຊື້ອສາຍແລະຄອບຄົວ. ມັນເກີດຂື້ນໃນ F plasmids ຜ່ານການຕ້ານເຊື້ອແບັກທີເຣຍ ວິທີຂອງການສືບພັນທາງເພດທີ່ເຫັນໃນແບັກທີເຣຍ. ໃນລະຫວ່າງການຕ້ານເຊື້ອແບັກທີເຣັຍ, R ປັດໄຈທີ່ມີ F plasmid ຕິດຕໍ່ກັບເຊື້ອແບັກທີເຣັຍອື່ນແລະການໂອນຍ້າຍ R ປັດໄຈລະຫວ່າງສອງເຊື້ອແບັກທີເຣັຍໂດຍຜ່ານການທົດລອງທາງເພດ. ແລະ, ນີ້ແມ່ນວິທີທົ່ວໄປທີ່ສຸດຂອງການແຜ່ກະຈາຍແລະການພັດທະນາຂອງການຕໍ່ຕ້ານຢາຕ້ານເຊື້ອໃນແບັກທີເຣຍ.

F Plasmid ແລະ R Plasmid ມີຄວາມຄ້າຍຄືກັນແນວໃດ?

  • F plasmid ແລະ R plasmid ແມ່ນສອງ plasmids. ເຊື້ອແບັກທີເຣັຍແມ່ນຈຸລິນຊີທີ່ບັນຈຸ plasmids ເຫຼົ່ານີ້. plasmids ຫຼາຍມີທັງປັດໄຈ F ແລະ R ປັດກັນ. ພວກມັນແມ່ນ DNA ພິເສດ. ທັງສອງປະເພດແມ່ນໂມເລກຸນ DNA ປິດ. ພວກມັນປະກອບດ້ວຍ DNA ທີ່ມີສອງແຖວ. ຍິ່ງໄປກວ່ານັ້ນ, ພວກມັນມີພັນທຸ ກຳ ທີ່ສະ ໜອງ ຄຸນລັກສະນະທີ່ເປັນປະໂຫຍດຕໍ່ເຊື້ອແບັກທີເຣັຍ. plasmids ເຫຼົ່ານີ້ສາມາດເຮັດແບບຕົວເອງໄດ້. ຍິ່ງໄປກວ່ານັ້ນ, ພວກມັນສາມາດແຜ່ເຊື້ອຈາກແບັກທີເຣຍ ໜຶ່ງ ໄປສູ່ອີກເຊື້ອສາຍ ໜຶ່ງ ແລະມີສ່ວນຮ່ວມໃນການໂອນ ກຳ ມະພັນທາງນອນ.

ຄວາມແຕກຕ່າງລະຫວ່າງ F Plasmid ແລະ R Plasmid ແມ່ນຫຍັງ?

F plasmid ແມ່ນ plasmid ທີ່ປະກອບດ້ວຍປັດໃຈການຈະເລີນພັນທີ່ ຈຳ ເປັນ ສຳ ລັບການມີເພດ ສຳ ພັນແລະການສ້າງເຊວເພດ ສຳ ພັນ. ໃນຂະນະດຽວກັນ R Plasmid ແມ່ນ plasmid ທີ່ບັນຈຸມີພັນທຸ ກຳ ທີ່ ຈຳ ເປັນ ສຳ ລັບການຕໍ່ຕ້ານກັບຢາຕ້ານເຊື້ອ. ດັ່ງນັ້ນ, ນີ້ແມ່ນຄວາມແຕກຕ່າງທີ່ ສຳ ຄັນລະຫວ່າງ F plasmid ແລະ R plasmid. ຍິ່ງໄປກວ່ານັ້ນ, plasmids F ແມ່ນມີຄວາມສາມາດໃນການປະກອບ pili. ໃນທາງກົງກັນຂ້າມ, plasmids R ທົ່ວໄປແມ່ນບໍ່ສາມາດຜະລິດ pili ທາງເພດໄດ້. ເພາະສະນັ້ນ, ນີ້ແມ່ນຄວາມແຕກຕ່າງອີກຢ່າງ ໜຶ່ງ ລະຫວ່າງ F plasmid ແລະ R plasmid.

ຍິ່ງໄປກວ່ານັ້ນ, ຄວາມແຕກຕ່າງທີ່ ສຳ ຄັນລະຫວ່າງ F plasmid ແລະ R plasmid ແມ່ນໄພຂົ່ມຂູ່ທີ່ພວກມັນກໍ່ໃຫ້ເກີດ. ນັ້ນ​ແມ່ນ ການແຜ່ລະບາດຂອງ F plasmid ບໍ່ເປັນໄພຂົ່ມຂູ່ທີ່ແທ້ຈິງເວັ້ນເສຍແຕ່ວ່າມັນມີປັດໄຈ R, ໃນຂະນະທີ່ການແຜ່ກະຈາຍຂອງ R plasmid ແມ່ນໄພຂົ່ມຂູ່ທີ່ແທ້ຈິງນັບຕັ້ງແຕ່ມັນຊ່ວຍໃນການພັດທະນາຄວາມຕ້ານທານຕໍ່ຢາຕ້ານເຊື້ອໃນປະຊາກອນແບັກທີເລຍ.

ສະຫຼຸບລວມ - F Plasmid vs R Plasmid

F plasmid ແມ່ນ plasmid ທີ່ເອົາປັດໃຈການຈະເລີນພັນທີ່ຊ່ວຍໃຫ້ການໂອນວັດຖຸພັນທຸ ກຳ ຈາກເຊື້ອແບັກທີເຣຍ ໜຶ່ງ ໄປສູ່ເຊື້ອແບັກທີເຣັຍອື່ນຜ່ານການເຊື່ອມຕົວເຂົ້າກັນ. ຍິ່ງໄປກວ່ານັ້ນ, plasmids F ແມ່ນ episomes ທີ່ສາມາດລວມ DNA ຂອງມັນເຂົ້າໄປໃນໂຄໂມໂຊມຂອງແບັກທີເຣຍອີກ. ໃນຂະນະທີ່, R plasmid ແມ່ນ plasmid ທີ່ມີປັດໃຈຕ້ານທານເຊິ່ງຊ່ວຍໃຫ້ການຕໍ່ຕ້ານກັບຢາຕ້ານເຊື້ອຫຼືຕົວຍັບຍັ້ງການຈະເລີນເຕີບໂຕຂອງແບັກທີເລຍ. plasmids ຫຼາຍມີທັງປັດໄຈ F ແລະ R ປັດໄຈ. ການແຜ່ກະຈາຍຂອງ R plasmids ແມ່ນໄພຂົ່ມຂູ່ທີ່ແທ້ຈິງກ່ວາການແຜ່ກະຈາຍຂອງ F plasmids ນັບຕັ້ງແຕ່ເຊື້ອແບັກທີເຣັຍໄດ້ຮັບການຕໍ່ຕ້ານກັບການປິ່ນປົວດ້ວຍຢາຕ້ານເຊື້ອ. ສະນັ້ນ, ນີ້ສະຫຼຸບຄວາມແຕກຕ່າງລະຫວ່າງ F plasmid ແລະ R plasmid.


1. “ ປັດໄຈ R-.” Wikipedia, ມູນນິທິວິກິພີເດຍ, ວັນທີ 28 ກັນຍາ 2019, ມີຢູ່ທີ່ນີ້. 2. “ Plasmids ແບັກທີເຣຍ: ຄຳ ນິຍາມ, ໜ້າ ທີ່ແລະການ ນຳ ໃຊ້.”,, ມີຢູ່ທີ່ນີ້.


1. “ Conjugation” ໂດຍ Adenosine - ວຽກງານຂອງຕົນເອງ (CC BY-SA 3.0) ຜ່ານ Commons Wikimedia 2. “ PBR322” ໂດຍ Ayacop (+ Yikrazuul) - ວຽກງານຂອງຕົນເອງ (ໂດເມນສາທາລະນະ) ຜ່ານ Commons Wikimedia


Resistance transfer factor (shortened as R-factor or RTF) is an old name for a plasmid that codes for antibiotic resistance. R-factor was first demonstrated in Shigella in 1959 by Japanese scientists. [1] Often, R-factors code for more than one antibiotic resistance factor: genes that encode resistance to unrelated antibiotics may be carried on a single R-factor, sometimes up to 8 different resistances. Many R-factors can pass from one bacterium to another through bacterial conjugation and are a common means by which antibiotic resistance spreads between bacterial species, genera and even families. [2] For example, RP1, a plasmid that encodes resistance to ampicillin, tetracycline and kanamycin originated in a species of Pseudomonas, from the family Pseudomonadaceae, but can also be maintained in bacteria belonging to the family Enterobacteriaceae, such as Escherichia coli. [3]

Bacteria containing F-factors (said to be "F+") have the capability for horizontal gene transfer they can construct a sex pilus, which emerges from the donor bacterium and ensnares the recipient bacterium, draws it in, and eventually triggers the formation of a mating bridge, merging the cytoplasms of two bacteria via a controlled pore. This pore allows the transfer of genetic material, such as a plasmid. Conjugation allows two bacteria, not necessarily from the same species, to transfer genetic material one way. [4] Since many R-factors contain F-plasmids, antibiotic resistance can be easily spread among a population of bacteria. Also, R-factors can be taken up by "DNA pumps" in their membranes via transformation, or less commonly through viral mediated transduction, or via bacteriophage, although conjugation is the most common means of antibiotic resistance spread. They contain the gene called RTF (Resistance transfer factor).

  1. ^ Macuch P, Seckarova A, Parrakova E, Krcmery V, Vymola F (1967). "Transfer of tetracycline resistance from Escherichia coli to other Enterobacteriaceae in vitro". Zeitschrift für Allgemeine Mikrobiologie. 7 (2): 159–62. doi:10.1002/jobm.19670070212. PMID4876865.
  2. ^
  3. Institute of Medicine (US) Forum on Microbial Threats (2010). "Workshop Overview: AMR Genes". Antibiotic Resistance: Implications for Global Health and Novel Intervention Strategies: Workshop Summary. National Academies Press. doi:10.17226/12925. ISBN978-0-309-18534-9 . PMID21595116. NBK54257/. The vast majority of antimicrobial resistance genes reside on mobile genetic elements such as insertion sequences, integrons, transposons, and plasmids, . Bacteria readily acquire these genetic elements from the environment, exchange them through conjugation, and receive them via infection by bacterial viruses (bacteriophages, or phages).
  4. ^
  5. Saunders, J. R. Grinsted, J. (November 1972). "Properties of RP4, an R Factor Which Originated in Pseudomonas aeruginosa S8". Journal of Bacteriology. 112 (2): 690–6. PMC251476 . PMID4628745.
  6. ^
  7. "Prokaryotic Cell Structure: Pili". Archived from the original on 2016-12-07 . Retrieved 2017-01-19 .

This molecular biology article is a stub. You can help Wikipedia by expanding it.

For the construction of plasmid DNA, in the beginning, the DNA of our interest is isolated from the genome.

Using the restriction digesting method followed by the PCR amplification, multiple copies of the gene of interest are generated.

Now the plasmid DNA is digested using the restriction endonuclease enzyme which creates the same sticky ends alike the gene of our interest.

Once it has done, along with the gene of interest, the promoter site (for performing replication) is inserted into the plasmid using the ligation method.

Enzyme ligase used to seal the ends of the plasmid DNA and insert (Ligase).

Once the process of plasmid construction is done, the plasmid DNA is amplified or can directly be inserted into the bacterial host.

In general, the E.coli is used for the transformation experiments.

The bacterial containing artificial plasmid is grown using the standard culturing procedure.

Once, the sufficient amount of bacterial colonies are cultured, the plasmid DNA is isolated.

Now the plasmid DNA is inserted into the target cell population in which we want to transfer our gene of interest.

(Or we can extract the therapeutic protein from it.)

There are many methods for gene transfer are available.

Electrophoration, sonication, liposome-mediated gene transfer and viral-vector- mediated gene transfer are some of them.

After the insertion, the plasmid DNA incorporated into our genome and replicated naturally.

This is the general outline of the use of plasmid DNA is gene transfer and gene therapy experiments.

The graphical illustration of the entire process of gene transfer or production of a therapeutic protein.

Biology Unit 4

2. Insertion of human DNA into plasmids
(D)igest (M)ix (F)orm
-Bacterial plasmid and cDNA of insulin gene are digested the same restriction enzyme- to produce complemetary sticky ends
Restriction enzyme cuts the plasmid DNA at its single restriction site, disrupting lac Z gene
-cDNA of insulin gene and the digested plasmids are mixed to allow sticky ends of DNA from both sources to anneal by complementary base pairing
-Mixture incubated with DNA ligase
Annealed strands are joined covalently by phosphodiester bonds, forming recombinant plasmid.
Ligation mixture will contain 3 major types of DNA molecules: reannealed plasmids, reannealed cDNA fragments and Recombinant plasmid

3. Transformation
- Cells are electroporated hence making pores appear transiently in the cell membranes of the bacterial cells, making the membrane permeable to DNA and allowing DNA molecules to enter the cells. (refer to fig 9 on p11)

4. Selection and Cloning of transformed cells with recombinant plasmids
-Transformed cells are placed onto nutrient agar plates containing the ampicillin and X-gal and incubated at 37°C
-Only bacteria transformed with the plasmid (reannealed and recombinant) will grow to form a colony as they have the amp^R gene that confers amplicilin resistance
-During formation of the recombinant plasmid, insertion of cDNA into the plasmid within the lac Z disrupted the lac Z gene
hence enzyme ß-galactosidase not produced
and X-gal not hydrolysed
thus bacterial colony carrying recombinant plasmid will appear White hence are picked since they contain recombinant plasmid with cDNA for insulin gene
-Bacterial colonies carrying reannealed plasmid will synthesised ß galactosidase which will hydrolyse the X-gal to produce blue pigment
-As bacteria multiply, plasmids within bacteria will multiply as well

5. Identification of bacterial colonies carrying gene of interest with Gene Probes
-White colonies containing recombinant plasmids are identified and grown on an agar plate and then transferred onto a filter
-DNA is then denatured with chemicals to separate the two strands
-radioactive DNA probe which is ssDNA complementary to a portion of the gene of interest is added to the filter. It will anneal to gene of interest by H-bonds
-Black spots formed on film during AUTORADIOGRAPHY show the positions of where the probe has bound to the gene of interest
-Based on the location of black spots, we can go to the original master plate to pick the colonies that are confirmed to have the gene of interest inserted into the plasmid.

(ALTERNATIVE METHOD OF SELECTING RECOMBINANTS: using 2 anitbiotic-resistant genes(amplicilin and Tet^R where restriction site lies within Tet^R)
(must refer to pg 14!)

Plasmid - Defination, Structure, Properties, replication, Copy number, Types, Functions and applications

Plasmid is a short, naturally occurring extra chromosomal, usually circular, double stranded DNA molecule that replicate, autonomously and lead an independent existence in Bacterial cell.

The term Plasmid was first given by Joshua Lederberg in 1952.

Properties of Plasmid :

• Plasmid is naturally found in cytoplasm, seperately from the main bacterial chromosome and are much smaller incomparision.

• Plasmids are mostly circular negatively supercoiled, double stranded DNA molecule. But linear Plasmids are also reported in genera streptomycetes and Borrelia. Also RNA plasmids are rare but reported in few plants, fungi and animal.

• Plasmids are normally in size range of 1kb to 250 kb which is smaller much smaller than Bacterial chromosome.
• Plasmids are considered as replicon. They replicate independently and code for their own transfer (i.e. Ori site present).
Smaller plasmid use host cell DNA replication machinary.
Larger plasmid carry gene for special enzymes specific for plasmid replication.

• Some are interogative plasmid called episomes. Also able to replicate by inserting them into bacterial chromosome and may be stably maintained in this form through numerous cell division, but show independent existence at some stage.

Copy number : Number of plasmids in an individual bacterial cell may very (1-100 or more) denoted by copy number.

• Plasmids are not essential for viability of bacterial cell. but genes carried by plasmids usually encodes traits beneficial for bacteria. e.g. antibiotic resistance, heavy metal resistance, Virulence factors, N2 fixation.

• Plasmid provide a mechanism of horizontal gene transfer via conjugation, transduction and transformation
Examples of plasmids : Puc 8 (E.cli), R-1, R-6, Col E1 (E.coli), Tol (Pseudomonas putida).

General Structure of Plasmid :

Structurel elements of bacterial plasmids may vary according to their size and function. Plasmid have following elements -

Structure of Plasmid

1. Origin of replication : DNA sequence which encode initiation of plasmid replication by recruiting bacterial transcription machinary for replication enzymes & proteins.

2. Antibiotic resistance gene : these genes provide a survival advantage to the bacterial host thet allows for selection of plasmid containing bacteria.

3. Multiple cloning site : Short segment containing a several restriction enzyme sites enabling easy insertion of foreign DNA.

4. Promoter region : It drives the transcription of the foreign insert.

5. Selectable marker : It is used to select for cells that has successfully taken up plasmid for the purpose of expression of inserted DNA.

6. Primer binding site : It is used as an initiation point for PCR amplification or sequencing of the plasmid.

Types of Plasmid (based on function) -

A. Fertility or F-plasmid :

B. Resistance or R-plasmid:

C. Col plasmid :

D. Metabolic/Degradative Plasmid :

E. Virulence Plasmid :

F. Suicide plasmid :

Types of plasmid based on their ability to transfer to another bacteria -

B. Non-conjugative plasmid -
This plasmid incapable of initiating conjugation, hence can only be transferred with the help of conjugative plasmid (tra-, mob-) under some circumstances.
E.g. many R plasmid, most Col plasmid.

C. Mobilisable plasmid :
This is an intermediate class of plasmid carry only a subset of genes (mob+) required for transfer.
They parasitize another plasmid, transferring at high frequency in presence of conjugative plasmid
(tra+, mob+).

Plasmid replication :

Plasmid are replicate autonomously and it also contain origin of replication site. There are two models are proposed for plasmid replication -
1) Theta θ model :
2) Rolling circle model :

1). Theta θ model :

Theta model replication of plasmid

2). Rolling circle model :

Host range :

Copy Number :

Copy number refers to the number of plasmid molecules of an individual.
Plasmid that are normally found in a single bacterial cell.
Plasmids are Classifying into two types based on copy no.
1). Stringent Plasmid :
- It is a large plasmid molecule.
- have low copy no. It present 1 to 2 per cell.
2). Relaxed plasmid :
- This plasmids are very small.
- have high copy no. 50 or more per cell.

Regulation of copy number :

Copy number is regulated by negative regulatory mechanisms and adjusting rate of plasmid replication.

1, by antisense RNA counter transcribes RNA,- plasmid replication is controlled by regulating the amount of available primer for replication.

2, Regulation by binding of replication proteins to introns (18-22 bp repeated units). Regulating amount of replication machinary available.

3, Regulation by ct RNA and protein, counter transcribed RNA limits the function of essential replication protein.

Plasmid incompatibility :

Incompatibility is defined as inability of two closely related plasmids to co-exist stably in the same host cell.
Several different kinds of plasmids may be found in a single cell, including more than one different conjugative plasmid.
To be able to coexist in the same cell, different plasmids must be compatible.
Different types of plasmids can therefore be assigned to different incompatibility groups, on the basis of whether or not they can coexist.

Incompatibility group : two plasmids that can not coexist in the same bacterial cell belong to a incompatibility group.

Why plasmids are incompatible ?

Partitioning :

It is active process that ensure that after cell division each daughter cell gets at least one copy of plasmid.
- Relaxed plasmid each daughter cell after division may get a copy by random diffusion or segregation.
- Stringent Plasmid most likely one daughter cell not received plasmid during segregation.

Par ABS system :- it is a broadly conserved molecular mechanisms for plasmid partitioning and chromosome segregation in bacteria.

Par ABS has 3 elements -
Par A - ATPas
Par B - DNA binding protein
Both are found on same operon.
Par S - cis acting sequence, located within or adjacent to this operon.

Confirmation of Plasmids :

Functions/Uses of plasmid (in genetic engineering) :

Advantages of using plasmid in genetic engineering :

Plasmid in eukaryotes :

- circular plasmid which is 6.3 kbcircular found in most Saccharomyces cerevisiae strain.
- 50-100 copies per haploid genome of yeast.
- This is the mostly studied eukaryotic plasmid.

Watch the video: What is a Plasmid? - Plasmids 101 (January 2023).