Segmented poly(a) tails

Abstract

The invention relates to a polynucleotide encoding or comprising a segmented poly(A) tail.

Description

[0001]Segmented poly(A) tails Field of the invention The present invention relates to a polynucleotide encoding or comprising a segmented poly(A) tail. The invention also relates to a vector comprising the polynucleotide, a cell comprising the polynucleotide or the vector, and a pharmaceutical composition comprising the RNA molecule. The invention further relates to a method of treating or preventing a disease using the pharmaceutical composition, a method of propagating the polynucleotide, a method of producing the RNA molecule, a method for producing a peptide, polypeptide or protein, and a method of increasing the stability or translational capacity of the polynucleotide encoding or comprising a poly(A) tail. Background of the invention The use of mRNA vaccination platforms in the SARS-CoV-2 pandemic transformed and accelerated the development of mRNA pharmaceuticals targeting a variety of other infectious diseases. The versatility, low cost and easily scalable production associated with this technology further sparked a renewed interest for its use in the realm of personalised medicine. mRNAs are currently designed to target cancers, reprogram the immune system and to provide therapies for rare disorders. The technology has the potential to provide new strategies to medicate currently incurable conditions or replace current costly treatments that are often associated with adverse reactions. mRNA production by in vitro transcription (IVT) is a relatively simple and a highly scalable chemical reaction. The process typically involves a single peptide DNA- dependent RNA polymerase originating from the T7 bacteriophage, ribonucleotides and a linearized plasmid DNA template. For in vitro transcribed mRNAs to function as blueprints for the translation of proteins in patient cells, they typically carry specific modifications at their 5’ and 3’ ends. In eukaryotic cells, mRNAs are made from primary transcripts that are co- transcriptionally processed to render them functional. This includes the modification of the very 5’ end of the RNA by capping. Capping is the addition of a methylated guanosine to the 5’ nucleotide that subsequently binds the nuclear cap binding complex. The cap structure is critical for the nuclear-cytoplasmic export of the mRNA and its stability. In addition, it is essential for efficient recruitment of ribosomes and the initiation of translation in the cytoplasm. Like the 5’ end, the 3’ end of almost all mRNAs is subject to a specific modification that involves a two-step reaction; cleavage and polyadenylation. The pre-mRNA, whilst attached to the elongating RNA polymerase II, is first cleaved at the so-called polyadenylation site and then subjected to the non-templated addition of around 200-250 adenosines. This process is catalysed by the poly(A) polymerase enzyme. The poly(A) tail in the nucleus is bound by the nuclear poly(A) binding protein (PABPN1) with a footprint of 11-14 adenosines. These polyadenylated and capped mRNAs are then exported from the nucleus to the cytoplasm where the cap and poly(A) binding proteins are exchanged. The nuclear cap binding proteins are replaced by the eukaryotic translation initiation multiprotein complex eIF4F. At the 3’ end of the mRNA, after exiting the nuclear pore, PABPN1 is replaced by its cytoplasmic counterpart PABPC1. Compared to PABPN1, PABPC1 has a larger footprint of approximately 24-27 adenosines (Baer & Kornberg, 1983; Sawazaki et al., 2018), although each pair of RNA recognition motif (RRM) domains within PABPC1 may bind to a stretch of as little as 11 adenosines (Deo et al., 1999). The current closed loop model suggests that the mRNA is circularised by the interaction between PABPC1 and eIF4G, a component of the eIF4F cytoplasmic cap binding complex. The circular mRNA is then recognised by ribosomal components and translation initiation commences. The poly(A) tail is tightly linked to the fate of the mRNA during its lifetime. It controls nuclear-cytoplasmic export, protects the mRNA from premature degradation by 3’-5’ exoribonucleases and is essential for efficient recruitment of the ribosome and translation initiation. The length of the poly(A) tails in newly synthesised mRNAs in the nucleus can reach up to 250 nucleotides but in the cytoplasm the tails of most mRNAs are trimmed to a median length of around 80-130 nucleotides in human cells. In tissue culture cells (HeLa / NIH3T3) poly(A) tails of cytoplasmic mRNAs are as short as 50-100 nucleotides. There is a complex relationship between the length of the poly(A) tail and the fate of endogenous mRNA in terms of its stability and the rate of translation. For example, in unfertilised animal eggs and early embryos, longer tailed mRNAs are translated with higher efficiency than shorter tailed mRNAs with little impact on stability. In tissue cultured Hela cells, translational rates are unaffected by poly(A) tail length, but length correlated with stability of the mRNA (Chang et al., 2014). There is much less ambiguity regarding poly(A) tail length and its link to translation and stability for exogenous mRNAs that are introduced into cells. In vitro transcribed (IVT) mRNAs introduced into host cells show a strong coupling between poly(A) tail length and translational output. A gradual increase in translational output is achieved by extending poly(A) tails up to 300 adenosines (Grier et al., 2016). Thus, to produce therapeutic mRNAs with maximal translational capacity, poly(A) tails are typically at least 100 nucleotides (Holtkamp et al., 2006) but ideally, reach up to 300 adenosines (Grier et al., 2016). The poly(A) tails in IVT mRNAs are template encoded and thus the plasmids that serve as templates harbour long adenosine homopolymer regions. However, it has been found that long adenosine homopolymers in circular plasmids are recombination hotspots that progressively shorten during plasmid amplification and / or subcloning processes (Grier et al., 2016; Trepotec et al., 2019). This plasmid instability represents a significant obstacle to maintain circular plasmids that can support in vitro transcription of mRNAs with long poly(A) tails. The use of mRNA vehicles in the realm of regenerative, replacement protein therapies, and expression of neutralising antibodies requires significantly higher expression of the target proteins compared to the levels of proteins that are sufficient to induce an effective immune reaction to combat viral infection (Rohner et al., 2022). To achieve this, mRNAs with long poly(A) tails that maximise translational output are required. This demands the design of novel poly(A) tail variants that can be incorporated into circular plasmids without triggering recombination that results in shortening of the template- encoded poly(A) tail during cloning and plasmid amplification steps. WO 2016 / 091391 A1 describes the use of a poly(A) tail comprising a first sequence of at least 60 adenine nucleotides and a second sequence of 20 to 400 adenine nucleotides. WO 2020 / 074642 A1 describes the use of a poly(A) tail comprising two segments of 60 adenine nucleotides, or three segments of 40 adenine nucleotides. WO 2016 / 005324 A1 describes the use of a poly(A) tail comprising 90 to 120 adenine nucleotides and a single spacer region. WO 2019 / 036513 A1 describes the use of a poly(A) tail comprising at least one homopolymer sequence of at least 8 consecutive adenine nucleotides and at least one interrupting sequence comprising one or more non-adenine nucleotides. An mRNA comprising a 165nt poly(A) tail is provided comprising a 3’ stretch of 63 consecutive adenosines (SEQ ID NO: 5), which does not show an improvement relative to an mRNA comprising a corresponding 101nt poly(A) tail lacking the 3’ stretch of 63 consecutive adenosines (SEQ ID NO: 4; Table 3). It is an aim of the present invention to improve the stability of sequences encoding a poly(A) tail, e.g. for use in IVT, and / or to improve translational output of mRNAs comprising a poly(A) tail, e.g. for therapeutic use. Summary of the invention The present inventors surprisingly found that a polynucleotide encoding a segmented poly(A) tail comprising at least 120 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22-35 consecutive A nucleotides, and wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides, or comprising a sequence of the formula AX(SAY)z, wherein S is a spacer sequence, X is 22-35, Y is 22-35 and Z is at least 4, and wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides, can be used to produce mRNAs having stable poly(A) tails of the desired length. As noted above, polynucleotides comprising sequences encoding conventional poly(A) tails may be susceptible to recombination. Thus, when they are produced (e.g. during cloning and propagation), the encoded poly(A) tails may be altered (e.g. shortened). Shorter poly(A) tails are generally not preferred as shortening the poly(A) tail of an mRNA may reduce the translational efficiency of the mRNA and / or its half-life. Unlike uninterrupted homopolymer adenosine stretches, the sequence encoding the segmented poly(A) tail is less susceptible to recombination, whilst an mRNA comprising the segmented poly(A) tail retains the translational efficiency and half-life of an mRNA comprising a conventional poly(A) tail. Thus, polynucleotides encoding such segmented poly(A) tails can be readily produced, for example, by propagation of a polynucleotide encoding the segmented poly(A) tail, and can serve as templates to transcribe mRNAs that can feature poly(A) tails of around 200 nucleotides or more, which may have increased translational output. As noted above, poly(A) tail length is linked to translation efficiency and stability in a cell type- and tissue specific-manner. Therefore, the present invention serves as a basis for a stable and versatile plasmid platform that supports the production of mRNAs with poly(A) tails of different lengths in order to optimise the efficacy of mRNA therapeutics. The invention is particularly suited to producing poly(A) tails of customizable length due to the ability to easily vary the number of stretches of consecutive A nucleotides within the poly(A) tract. This is a major advantage over other poly(A) tail designs that stabilise the poly(A) tract in E. coli, such as BioNTech’s 110nt split poly(A) tract (Vogel et al., 2021). Accordingly, the invention provides a polynucleotide comprising a nucleic acid sequence encoding a segmented poly(A) tail, wherein the segmented poly(A) tail comprises at least 120 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22-35 consecutive A nucleotides, and wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides. The invention also provides a polynucleotide comprising a nucleic acid sequence comprising a segmented poly(A) tail, wherein the polynucleotide is an RNA molecule, wherein the segmented poly(A) tail comprises at least 120 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22-35 consecutive A nucleotides, and wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides. The invention further provides a polynucleotide comprising a nucleic acid sequence encoding a segmented poly(A) tail, wherein the segmented poly(A) tail comprises a sequence of the formula AX(SAY)z, wherein S is a spacer sequence, X is 22-35, Y is 22- 35 and Z is at least 4, and wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides. The invention further provides a polynucleotide comprising a nucleic acid sequence comprising a segmented poly(A) tail, wherein the polynucleotide is an RNA molecule, wherein the segmented poly(A) tail comprises a sequence of the formula AX(SAY)z, wherein S is a spacer sequence, X is 22-35, Y is 22-35 and Z is at least 4, and wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides. The invention further provides a vector comprising the polynucleotide of the invention. The invention further provides a cell comprising the polynucleotide or the vector of the invention. The invention additionally provides an RNA molecule obtained or obtainable by transcription of, or encoded by, the polynucleotide or the vector of the invention. The invention also provides a pharmaceutical composition comprising the polynucleotide or RNA molecule of the invention. The invention also provides the pharmaceutical composition of the invention for use in a method of treating or preventing a disease in a subject. The invention also provides a method of propagating a polynucleotide, the method comprising propagating the polynucleotide or the vector of the invention. The invention also provides an in vitro method of producing an RNA molecule, the method comprising contacting the polynucleotide of the invention, the vector of the invention, the cell of the invention, or the propagated polynucleotide obtained by the method of propagating a polynucleotide of the invention, with an RNA polymerase. The invention also provides a pharmaceutical composition produced by the method of producing an RNA molecule of the invention, wherein said method further comprises formulating the RNA molecule into a pharmaceutical composition. The invention further provides an in vitro or in vivo method of producing a peptide, polypeptide or protein, the method comprising translating the polynucleotide or RNA molecule of the invention, or obtained by the method of producing an RNA molecule of the invention. The invention additionally provides a method of increasing the stability or translational capacity of a polynucleotide comprising a nucleic acid sequence encoding or comprising a poly(A) tail, comprising replacing the sequence encoding or comprising the poly(A) tail with a sequence encoding or comprising a segmented poly(A) tail as defined in the polynucleotide of the invention. Brief description of the figures Figure 1 - Plasmids featuring a 120-nucleotide-long adenosine homopolymer are unstable in bacterial hosts. The plasmid encoding a 120-nucleotide-long homopolymer adenosine tract undergoes recombination during subcloning resulting in daughter clones (subclones 1 – 3) with shortened polyadenosine tracts. The features of the plasmids are indicated, 5’UTR: 5’ untranslated region, 3’UTR: 3’ untranslated region, ORF: open reading frame. The location of the KpnI and SapI restriction enzyme sites flanking the templated adenosine stretch AAAAAnare shown. These clones where the ORF was successfully exchanged all featured shortened poly(A) tracts ranging from 30 – 95 adenosines as determined by Sanger sequencing. Figure 2 – GA27-segmented poly(A) tail design. The GA27poly(A) segment approach allows the construction of plasmids that feature poly(A) tracts with different lengths including longer than 120 nucleotides. Figure 3 - GA27-segmented poly(A) design stabilises poly(A) tract during cloning to allow production of extremely long poly(A) tracts. The poly(A) tracts of plasmids from 4 randomly picked clones featuring 120 uninterrupted adenosines, or a 114 or 198 nucleotide long GA27-segmented poly(A) tracts were assessed for shortening by sequencing (shortening increases the number of non-A nucleotides). For all clones, the number of each of the four sequenced nucleotides are represented by different textured bars with the number of adenosines represented by filled black bars. N bases are those that have not been assigned as A, T, G, or C by the sequencing algorithm. Figure 4 –Plasmids featuring the GA27-segmented poly(A) design or the BioNTech split poly(A) design have superior stability compared to uninterrupted poly(A) tracts of similar length. After cloning each distinct poly(A) tract from the same parental vector DNA containing a 30nt poly(A) tract, plasmid DNA was purified from individual clones and digested with BssHII and HindIII. As these restriction enzymes excise the 3’UTR and poly(A) tract, the differential migration of the band reflects the length of the poly(A) tract. Digestion of plasmids containing an uninterrupted 120nt poly(A) tract (A) is expected to excise a 439bp fragment, whereas digestion of plasmids with 114nt GA27-segmented poly(A) tract (B) is expected to excise a 433bp fragment. Digestion of plasmids with the 110nt BioNTech split poly(A) tract (C) is expected to excise a 429bp fragment. Black arrow (marking the upper band in (A) clone 9, (B) clone 4 and (C) clone 8) indicates the position of the restriction fragment containing an expected full-length polyadenosine tract, white arrow (marking the lower band in (A) clone 9, (B) clone 4 and (C) clone 8) indicates fragments with a shortened polyadenosine tract. The sizes of the most relevant marker bands are indicated beside the gels. Quick- Load 1kb plus DNA ladder was loaded in each DNA ladder lane (L). Figure 5 – The GA27-segmented poly(A) tail design does not compromise the translational capacity of the mRNAs. A) Schematic illustrating in vitro transcription of an mRNA featuring a 114-nucleotide-long GA27-segmented poly(A) tail using a SapI- linearised plasmid as the template. B) Comparison of the translational capacity of mRNAs that either feature an uninterrupted 30nt or 120nt long poly(A) tail or a 114nt GA27-segmented poly(A) tail in HEK-293T cells. 1μg of each of the different luciferase- encoding mRNAs was co-transfected with the same amount of monoclonal antibody (R5.016)-encoding mRNAs (Alanine et al., 2019) for normalization. Translational capacity is calculated by dividing luciferase activity in each well by R5.016 concentration from the same well. At 48 hours post-transfection, the media is removed and R5.016 concentration is determined by indirect ELISA. After removing the media, cells are lysed and assayed for luciferase activity. Data is represented as mean ± SD.3 biological repeats were performed for each transfection condition. Figure 6 - The translational capacity of mRNAs featuring the GA27-segmented poly(A) design is at least as high as those with the BioNTech split poly(A) tail. Comparison of the translational capacity of mRNAs that either feature an uninterrupted 30nt or 120nt long poly(A) tail, a GA27-segmented poly(A) tail of 114nt or 198nt in length, or BioNTech’s 110nt long split poly(A) tail (Vogel et al., 2021). 1μg of each of the different luciferase-encoding mRNAs was co-transfected into HEK-293T cells with monoclonal antibody (R5.016)-encoding mRNAs (Alanine et al., 2019) for normalisation. Translational capacity is calculated by dividing luciferase activity in each well by R5.016 concentration from the same well. At 48 hours post-transfection, the media is removed and R5.016 concentration is determined by indirect ELISA. After removing the media, cells were lysed and assayed for luciferase activity. Data is represented as mean ± SD (2 biological repeats for mRNA containing a 30nt uninterrupted poly(A) tail and 3 biological repeats for all other mRNAs). Figure 7 – The sequence integrity of the GA27poly(A) segments is readily identified by Sanger sequencing. Sanger sequencing of the Renilla luciferase-encoding plasmid with the 114 nucleotide long GA27-segmented poly(A) tract. The guanosine nucleotide peaks between the adenosine stretches are clearly recognisable and demarcate the individual GA27segments, simplifying quality control and verification of plasmid integrity by Sanger sequencing. Sanger sequencing spectra is autoscaled. Figure 8 – Plasmids featuring a 114nt long GA27-segmented poly(A) tract are stable during amplification in E. coli. Quality control of the poly(A) tract region in Renilla luciferase-encoding plasmids purified from 3 mL (miniprep = Mini) or 100 mL (midiprep = Midi) E. coli overnight cultures by restriction enzyme digest and gel electrophoresis, and by Sanger sequencing. All plasmids are identical apart from the poly(A) tract sequence, which consisted of an uninterrupted 120nt poly(A) tract (A), a 114nt GA27-segmented poly(A) tract (B) or the BioNTech 110nt split poly(A) tract (C). A-C, miniprep cultures were used to inoculate 100 mL overnight cultures to prepare midiprep DNA plasmids. Restriction enzymes (BssHII and HindIII) were used to excise a DNA fragment containing the 3’UTR and poly(A) tract. Plasmids featuring an uninterrupted poly(A) tail show a smear (diffuse band) below the expected size representing plasmids with shortened poly(A) tracts (A). The Sanger sequencing spectra of the poly(A) region in each plasmid preparation is displayed below each respective digest lane. Figure 9 –Plasmids featuring 114nt GA27-segmented poly(A) tracts are stable during re-transformation and amplification in E. coli. The Renilla luciferase-encoding plasmids used to transform E. coli were purified from two separate clones (clone 2 and clone 3), both containing verified 114nt GA27- segmented poly(A) tracts. Miniprep plasmid DNA from these two clones was re- transformed and plasmids from five daughter clones each were analysed by restriction enzyme digest (BssHII and HindIII) to verify poly(A) tract length integrity. Plasmids with a fully intact 114nt GA27-segmented poly(A) tract excise a 433bp fragment (black arrow). White arrow indicates position of DNA fragments containing a shortened poly(A) tract. Quick-Load 1kb plus DNA ladder was loaded in each DNA ladder lane (L). The sizes of the most relevant marker bands are indicated to the right of the gel. Figure 10 - GA27 segments reduce susceptibility to shortening of the poly(A) tract during subcloning of IVT plasmids. Central gel depicts a control digest of the parental plasmids that were used to assess the stability of the different poly(A) tract designs during subcloning. Poly(A) tract integrity of the parental clone and its subclones is assessed by restriction enzyme digests. The gel on the left shows the analysis of subcloned plasmids featuring the GA27-segmented poly(A) tracts. The gel on the right shows analysis of subcloned plasmids using a parental clone that features a 120nt long uninterrupted poly(A) tract. Sanger sequencing of the clones confirms the lengths of the poly(A) tract in each plasmid as indicated below the gel. Digestion of plasmids containing an uninterrupted 120nt poly(A) tract (gel on the right) is expected to excise a 439bp fragment, whereas digestion of plasmids with 114nt GA27-segmented poly(A) tract (gel on the left) is expected to excise a 433bp fragment. Black arrow (marking the upper band of the 3’UTR + poly(A) tract in left and right panels) indicates the position of the restriction fragment containing an expected full-length polyadenosine tract, white arrow (marking the lower band of the 3’UTR + poly(A) tract in left and right panels) indicates fragments with a shortened polyadenosine tract. Quick-Load 1kb plus DNA ladder was loaded in each DNA ladder lane (L). The sizes of the most relevant marker bands are indicated by the gels. Figure 11 - Evaluating the stability of BioNTech’s 110nt split poly(A) tract during subcloning. Left gel confirms the integrity of the parental plasmid used for subcloning stability analysis. For subcloning, the Luciferase ORF was excised and replaced with an alternative, open reading frame 1 (ORF1). The ligated plasmids were transformed and plasmids from four randomly chosen subclones were analyzed by restriction enzyme digests excising fragments that confirm ORF1 inclusion and determine poly(A) tract lengths (3’UTR+poly(A) tract). Digestion of plasmids with the 110nt BioNTech split poly(A) tract is expected to excise a 429bp fragment (black arrow; the upper band of the 3’UTR + poly(A) tract in lane 4 of right panel), whereas white arrow (marking the lower band of the 3’UTR + poly(A) tract in lane 4 of right panel) indicates fragments with a shortened polyadenosine tract. The length of the poly(A) tracts were measured by Sanger sequencing. Quick-Load 1kb plus DNA ladder was loaded in each DNA ladder lane (L). The sizes of the most relevant marker bands are indicated to the right of each gel. Figure 12 – Plasmids featuring 198nt GA27-segmented poly(A) tracts can be cloned but are less stable during subcloning. Left gel shows a digest of the parental plasmid with a 198-nucleotide-long GA27-segmented poly(A) tract used for subcloning (dashed box). After subcloning to replace the luciferase ORF with ORF1, plasmids were purified and digested with restriction enzymes that excise ORF1 and a DNA fragment containing both the 3’UTR and poly(A) tract. Approximate lengths of the poly(A) tract region determined by Sanger sequencing are indicated below their respective gel lanes. Black arrow indicates the position of the restriction fragment containing an expected full-length polyadenosine tract (198nt poly(A) tract: 517 bp; marking the upper band of the 3’UTR+poly(A) tract in clone 1 of right panel), white arrows (marking the bands of the 3’UTR+poly(A) tract in clones 2 and 3 of right panel) indicate fragments with shortened polyadenosine tracts. Quick-Load 1kb plus DNA ladder was loaded in each DNA ladder lane (L). The sizes of the most relevant marker bands are indicated to the left of the gels. Figure 13 – If shortening of the 114nt GA27-segmented poly(A) tract occurs during subcloning, entire GA27 segments are lost, simplifying quality control. After subcloning to replace the Renilla luciferase (R. luc) ORF in the cloning vector (V, lane 2) with ORF2, plasmids were purified from 8 subclones and digested with restriction enzymes (HindIII, XhoI and BssHII) that excise ORF2 and a DNA fragment containing both the 3’UTR and poly(A) tract. Digestion of plasmids with a 114nt GA27-segmented poly(A) tract is expected to excise a 433bp fragment, the position of which is marked on the left of the gel. Sanger sequencing spectra for the cloning vector and subclones 2,4 and 8 are shown below respective digest lanes. Quick-Load 1kb plus DNA ladder was loaded in each DNA ladder lane (L). The sizes of the most relevant marker bands are indicated. Figure 14 - Production of in vitro transcription plasmids featuring poly(A) tracts of customizable length. A) Plasmid DNA purified from bacterial cultures (midiprep) digested with XhoI, HindIII and BssHII excising the ORF, and the 3’UTR and poly(A) tract. Digestion of plasmids containing an uninterrupted 30nt poly(A) tract is expected to excise a 358bp fragment, digestion of plasmids with the 110nt BioNTech split poly(A) tract is expected to excise a 429bp fragment. Digestion of plasmids featuring a 114nt or 198nt GA27-segmented poly(A) tract is expected to excise a 433bp fragment or 517bp fragment, respectively. Quick-Load 1kb plus DNA ladder was loaded in each DNA ladder lane (L). The sizes of the most relevant marker bands are indicated. B) as in A), but including plasmids featuring GA27-segmented poly(A) tracts ranging from 58 to 198 nucleotides. Figure 15 – Length of 198nt GA27-segmented poly(A) tract can be maintained during subcloning. Gel electrophoresis of ten 198nt GA27-segmented poly(A) plasmid subclones digested with XhoI, HindIII and BssHII. V, digestion of original plasmid used to create the vector to replace the Renilla luciferase ORF with ORF3. Position of the ORF3 fragment (expected size 1670bp), the Renilla luciferase ORF fragment (expected size 959bp) and the 198nt GA27segmented poly(A) + 3’UTR fragment (expected size 517bp) are indicated to left of the gel. Quick-Load 1kb plus DNA ladder was loaded in each DNA ladder lane (L). The sizes of the most relevant marker bands are indicated to the right of the gel. Figure 16 – Stability of a 198nt long GA27-segmented poly(A) tract during amplification in E. coli. Gel electrophoresis of XhoI, HindIII and BssHII digested mini and midiprep plasmids featuring two different lengths ORFs (ORF3 and ORF4) following amplification in E. coli. Quick-Load 1kb plus DNA ladder was loaded in each DNA ladder lane (L). The sizes of the most relevant marker bands are indicated and the position of the excised ORFs and GA27 segmented poly(A) + 3’UTR fragment are indicated. Figure 17 - Translational output of 170nt and 198nt GA27-segmented poly(A) tails exceeds other poly(A) designs. A) Gel electrophoresis of in vitro transcribed Renilla luciferase-encoding mRNAs featuring a 30nt uninterrupted poly(A) tail, the BioNTech poly(A) design or GA27-segmented poly(A) tails of 114, 170 or 198 nucleotides. B) Comparison of the luciferase activity obtained from mRNAs featuring the different poly(A) tails transfected into HEK-293T cells (48h). Data is represented as mean ± SD.3 biological repeats were performed for each mRNA variant. Figure 18 – Translational output of mRNAs with customizable length of GA27- segmented poly(A) tails. A) Gel electrophoresis of in vitro transcribed Renilla luciferase-encoding mRNAs featuring a 30nt uninterrupted poly(A) tail, or GA27- segmented poly(A) tails of 58, 86, 114, 142, 170 or 198 nucleotides. B) Comparison of the translational capacity of Renilla luciferase-encoding mRNAs (HEK-293T, 48h) that feature poly(A) tails ranging from 30 to 198 nucleotides in length. Figure 19 - Translational output of mRNAs with 114nt or 198nt poly(A) tail at different amounts. Comparison of the translational output from Renilla luciferase- encoding mRNAs that feature a 114nt or 198nt GA27-segmented poly(A) tails, at amounts of 1000ng, 750ng or 500ng. Data is represented as mean ± SD.3 biological repeats were performed for each transfection condition. Detailed description General definitions Unless defined otherwise, technical and scientific terms used herein have the same meaning as commonly understood by a person skilled in the art to which this invention belongs. In general, the term “comprising” is intended to mean including but not limited to. For example, the phrase “a pharmaceutical composition comprising an RNA molecule” should be interpreted to mean that the pharmaceutical composition comprises the RNA molecule, but that the pharmaceutical composition may comprise further components (for example an excipient, a lipid nanoparticle, In some embodiments of the invention, the word “comprising” is replaced with the phrase “consisting of”. The term “consisting of” is intended to be limiting. For example, the phrase “a pharmaceutical composition consisting of an RNA molecule” should be understood to mean that the pharmaceutical composition contains the RNA molecule and no further components. In some embodiments of the invention, the word “comprising” is replaced with the phrase “consisting essentially of”. The term “consisting essentially of” means that specific further components can be present, namely those not materially affecting the essential characteristics of the subject matter. For example, the phrase “a pharmaceutical composition consisting essentially of an RNA molecule” indicates that the pharmaceutical composition may further comprise one or more excipients that have no particular function. The term “about” or “around” when referring to a value refers to that value but within a reasonable degree of scientific error. Optionally, a value is “about X” or “around X” if it is within 10%, within 5% or within 1% of X. The singular forms “a”, “an” and “the” include plural references unless the context clearly dictates otherwise. All publications, patents and patent applications cited herein, whether Supra or Infra, are hereby incorporated by reference in their entirety. Polynucleotides The present invention relates to a polynucleotide comprising a nucleic acid sequence encoding or comprising a segmented poly(A) tail. The polynucleotide may be a single-stranded or a double-stranded polynucleotide. The polynucleotide may comprise a sense strand (otherwise known as a non-template strand or coding strand) and / or an antisense strand (otherwise known as a template strand or non-coding strand upon which the polymerase acts). Sense and antisense strands are typically complementary. The term “encoding a segmented poly(A) tail” is intended to mean that the nucleic acid sequence comprises sequence information from which the segmented poly(A) tail may be derived. A feature is therefore “encoded” by the nucleic acid sequence when the nucleic acid sequence comprises a sequence corresponding to the feature or its reverse complement. In an embodiment wherein the feature encoded is an RNA molecule and the nucleic acid sequence is a DNA molecule, the DNA nucleotides A, C, G and T correspond to the RNA nucleotides A, C, G and U, respectively, or their reverse complement U, G, C, A, respectively. For example, where the “segmented poly(A) tail” refers to the sequence A30GA27GA27, a nucleic acid sequence encoding the segmented poly(A) tail may comprise the sequence A30GA27GA27on a sense strand, and / or may comprise the sequence T27CT27CT30on an antisense strand. The term nucleic acid sequence is typically a transcribable nucleic acid sequence. The term “transcribable” means that the nucleic acid sequence in the polynucleotide, or its reverse complement, is capable of being transcribed, for example, by contact with a polymerase, such as a DNA-dependent RNA polymerase or an RNA-dependent RNA polymerase, to produce a further polynucleotide, such as an RNA molecule. In some cases, the invention relates to a polynucleotide comprising a nucleic acid sequence comprising a segmented poly(A) tail, wherein the polynucleotide is an RNA molecule. In other words, the invention relates to an RNA polynucleotide comprising a nucleic acid sequence comprising a segmented poly(A) tail. The segmented poly(A) tail comprises consecutive adenosine (A) nucleotides separated by two or more spacer sequences, to thereby form distinct segments of polyadenosine (poly(A)). A sequence of consecutive A nucleotides may be referred to herein as a “stretch” of A nucleotides. The segmented poly(A) tail, when present at the 3’ end of a messenger RNA (mRNA) transcript, may for example, bind to multiple molecules of poly-A binding protein C1 (PABPC1) and / or circularise (for example via the eukaryotic initiation factors eIF-4E and eIF-4G). The segmented poly(A) tail comprises stretches of at least 11 A nucleotides, typically 11 to 35 A nucleotides, separated by the spacer sequences. Without being bound by theory, the minimal binding footprint of PABPC1 is understood to be approximately at least 11 A nucleotides, or at least 22 A nucleotides for the binding footprint of all RRM domains in PABPC1, and typically extends to around 27 to 30 A nucleotides. Thus, a stretch of 11 to 35 consecutive A nucleotides is approximately the same length as the binding footprint of PABPC1, and a segmented poly(A) tail comprising such stretches may retain the ability of a conventional, non- segmented poly(A) tail to bind PABPC1, thus permitting circularisation of mRNA molecules. Furthermore, binding of PABPC1 and eukaryotic initiation factors is believed to reduce and / or prevent access of exoribonucleases, thus preventing the degradation of the mRNA molecule. The similarity between the length of the poly(A) stretch and the binding footprint of PABPC1 may further reduce the presence of unbound poly(A) overhangs that could be targeted by deadenylation enzymes. Accordingly, the segmented poly(A) tail has a number of advantages over conventional poly(A) tails in terms of stability in the cell. Furthermore, the use of two or more spacer sequences improves the stability of the nucleic acid sequence encoding the segmented poly(A) tail. As discussed herein, interruption of the sequence encoding the poly(A) tail with spacer sequences reduces of its susceptibility to recombination events occurring during cloning and propagation, when compared to sequences encoding a poly(A) tail with no spacer sequences and / or spacer sequences that are further apart. The sequence encoding the segmented poly(A) tail in the polynucleotide may exhibit higher stability than a control polynucleotide encoding a non-segmented poly(A) tail comprising the same number of A nucleotides. The stability may be calculated following propagation of the polynucleotide in E. coli. The control polynucleotide is identical to the polynucleotide of the invention except in the sequence of the poly(A) tail. For example, if the segmented poly(A) tail comprises the sequence A30GA27GA27GA27, the poly(A) tail of the control polynucleotide comprises the sequence A111. The term “higher stability” is intended to refer to the increased probability that the length of the segmented poly(A) tail is unchanged before and after the propagation when compared to the poly(A) tail of the control polynucleotide. Propagation of the polynucleotide may be performed by transforming E. coli with the polynucleotide, and subsequently culturing the E. coli under conditions suitable for increasing the number of molecules of the polynucleotide, for example by increasing the copy number per cell and / or increasing the number of cells. The transformation may be performed by heat shock. The step of culturing the E. coli may be performed for 6, 12, 18, 24 or 48 hours, for example, at 30˚C. The step of culturing the E. coli may be performed for 6, 12, 18, 24 or 48 hours, for example, at 37˚C. The culture is typically performed by plating the E. coli (e.g. on LB agar) following transformation. The culture may be performed under conditions to select for E. coli comprising the polynucleotide over E. coli not comprising the polynucleotide. The stability may be calculated as a percentage of the polynucleotides following propagation that retain the original sequence of the poly(A) tail. The sequence encoding the segmented poly(A) tail in the polynucleotide may exhibit at least 10 % higher stability when compared to the control polynucleotide, such as at least 20 %, at least 30 %, at least 40 %, at least 50 %, at least 60 %, at least 70 %, at least 80 %, at least 90 %, at least 100 %, at least 150 % or at least 200 % higher stability when compared to the control polynucleotide. The sequence encoding the segmented poly(A) tail in the polynucleotide may exhibit on average at least 10 % higher stability when compared to the control polynucleotide, such as on average at least 20 %, at least 30 %, at least 40 %, at least 50 %, at least 60 %, at least 70 %, at least 80 %, at least 90 %, at least 100 %, at least 150 % or at least 200 % higher stability when compared to the control polynucleotide. For example, if 25 % of the sequences encoding the non-segmented poly(A) tail in the control polynucleotide retain the original sequence following propagation, but 75 % of the sequences retain the original sequence in the polynucleotide of the invention, this is considered to be 200 % higher stability than the control polynucleotide. The length of the segmented poly(A) tail may be determined by any suitable means, for example, by DNA sequencing of E. coli clones following propagation and / or digestion of the plasmid, PCR amplification and gel electrophoresis to determine the size of a fragment containing the poly(A) tail in an E. coli clone following propagation. The sequence encoding the segmented poly(A) tail in the polynucleotide typically exhibits reduced propensity for recombination than a control polynucleotide encoding a non-segmented poly(A) tail comprising the same number of A nucleotides, typically calculated following propagation of the polynucleotide in E. coli. The control polynucleotide is identical to the polynucleotide of the invention except in the sequence of the poly(A) tail. For example, if the segmented poly(A) tail comprises the sequence A30GA27GA27GA27, the poly(A) tail of the control polynucleotide comprises the sequence A111. The term “reduced recombination” is intended to refer to the reduced probability that the sequence of the segmented poly(A) tail undergoes recombination during propagation when compared to the poly(A) tail of the control polynucleotide. Without being bound by theory, it is understood that the recombination that occurs in non-segmented poly(A) tails is due to homologous recombination of similar or identical nucleotide sequences, i.e. poly(A), and the presence of spacer sequences reduces the frequency and positions at which homologous recombination may occur. Propagation of the polynucleotide may be performed by transforming E. coli with the polynucleotide, and subsequently culturing the E. coli under conditions suitable for increasing the number of molecules of the polynucleotide, for example by increasing the copy number per cell and / or increasing the number of cells. The recombination may be calculated as a percentage of the polynucleotides following propagation that retain the original sequence of the poly(A) tail. Recombination may be reduced by at least 5 % when compared to the control polynucleotide, such as at least 10 %, at least 20 %, at least 30 %, at least 40 %, at least 50 %, at least 60 %, at least 70 %, at least 80 %, at least 90 % or at least 95 % when compared to the control polynucleotide. The sequence of the segmented poly(A) tail may be determined by any suitable means, for example, by DNA sequencing of E. coli clones following propagation and / or digestion of the plasmid, PCR amplification and gel electrophoresis to determine the size of a fragment containing the poly(A) tail in an E. coli clone following propagation. The nucleic acid sequence may encode an RNA molecule having the segmented poly(A) tail, and the RNA molecule may have the same or greater translational capacity than a control RNA molecule comprising a non-segmented poly(A) tail comprising the same number of A nucleotides. The RNA polynucleotide comprising a nucleic acid comprising the segmented poly(A) tail may have the same or greater translational capacity than a control RNA molecule comprising a non-segmented poly(A) tail comprising the same number of A nucleotides. The control RNA molecule is identical to the RNA molecule encoded by the polynucleotide of the invention or the RNA polynucleotide of the invention except in the sequence of the poly(A) tail. For example, if the segmented poly(A) tail comprises the sequence A30GA27GA27GA27, the poly(A) tail of the control RNA molecule comprises the sequence A111. The term “translational capacity” is intended to refer to the amount of polypeptides translated from the RNA molecule over a specific period of time. Without being bound by theory, the translational capacity of an RNA molecule may be affected by the stability of the poly(A) tail within a cell, such that shorter, non-segmented tails may degrade more quickly and thus will be unable to serve as a template for translation, whilst poly(A) tails that are unable to circularise with a 5’mRNA cap may lead to reduced translation due to slower binding by ribosomes. Furthermore, RNA molecules with a shorter poly(A) tail may be less competitive for the pool of PABPC1 molecules within a cell than an RNA molecule with a longer poly(A) tail having many PABPC1 binding sites, and so may spend more time without any PABPC1 coating its poly(A) tail and resulting in reduced circularisation and translation. For the purposes of determining translational capacity, the RNA molecule and the control RNA molecule may be provided as part of a larger nucleic acid sequence each comprising a sequence encoding a luciferase, such as a Renilla luciferase. The sequence encoding the luciferase may be upstream, i.e.5’ of, the segmented poly(A) tail, such as immediately upstream of the segmented poly(A) tail. The translational capacity may be calculated as the measured level of luciferase expressed 48 hours after transformation of the RNA molecule into a HEK293T host cell. The measured level may be an absolute level, or may be a relative level, e.g. the level produced by the RNA molecule relative to the control RNA molecule. The level may be measured by any means known to the skilled person, for example, in a luciferase activity assay in the presence of the substrate luciferin and excess ATP, or by quantitative mass-spectrometry. Translational capacity may be considered to be the ‘same’ when the level of luciferase expressed from the RNA molecule is within 20% of the level of luciferase expressed from the control RNA molecule, such as within 10% or within 5 %. Translational capacity may be considered to be the ‘greater’ when the level of luciferase expressed from the RNA molecule is at least 20% more, at least 30% more, at least 40% more or at least 50% more. The nucleic acid sequence may encode an RNA molecule having the segmented poly(A) tail, and the RNA molecule may have the same or greater half-life than a control RNA molecule comprising a non-segmented poly(A) tail comprising the same number of A nucleotides. The RNA polynucleotide comprising a nucleic acid comprising the segmented poly(A) tail may have the same or greater half-life than a control RNA molecule comprising a non-segmented poly(A) tail comprising the same number of A nucleotides. The control RNA molecule is identical to the RNA molecule encoded by the polynucleotide of the invention or the RNA polynucleotide of the invention except in the sequence of the poly(A) tail. For example, if the segmented poly(A) tail comprises the sequence A30GA27GA27GA27, the poly(A) tail of the control RNA molecule comprises the sequence A111. The term “half-life” is intended to refer to the time taken for 50 % of the RNA molecules to be degraded such that they can no longer express a full amino acid sequence encoded by the RNA molecule. Without being bound by theory, the segmented nature of the poly(A) tail, together with the specifically designed lengths of the stretches of consecutive A nucleotides reduces the action of deadenylation enzymes. Furthermore, the segmented nature of the poly(A) tail described herein allows for the production of much longer poly(A) tails which require more deadenylation reactions to be performed until the RNA molecule is degraded to the extent that it can no longer express a full amino acid sequence encoded by the RNA molecule. For the purposes of determining half-life, the RNA molecule and the control RNA molecule may each comprise a sequence encoding a luciferase, such as a Renilla luciferase. The half- life may be calculated by any means known in the art. For example, the half-life may be calculated by Northern blot, RT-PCR, fluorescent hybridisation (e.g. nanostring) or quantitative RNA-sequencing. The half-life may be calculated at periodic intervals following transformation of the RNA molecule into a HEK293T host cell. The half-life may be considered to be the ‘same’ when the half-life of the RNA molecule is within 20% of the half-life of the control RNA molecule, such as within 10% or within 5 %. The half-life may be considered to be the ‘greater’ when the half-life of the RNA molecule is at least 20% more, at least 30% more, at least 40% more or at least 50% more. The polynucleotide may be a DNA polynucleotide or an RNA polynucleotide. The DNA polynucleotide may be a single-stranded or double-stranded DNA polynucleotide. The RNA polynucleotide may be a single-stranded or double-stranded DNA polynucleotide. Polynucleotides have a chemical orientation defined by the position of the linking carbon in the five-carbon sugar of each consecutive nucleotide in the chain. Accordingly, sequence elements positioned sequentially along the length of a polynucleotide may be defined by the directionality of the chain of nucleotides that is either 5’ to 3’ or 3’ to 5’. DNA polynucleotides typically comprise a combination of adenosine (A), guanosine (G), cytidine (C) and thymidine (T) nucleotides. In an RNA polynucleotide, the T nucleotides are typically replaced by uridine (U), which retains the ability to base pair with A. RNA polynucleotides therefore typically comprise a combination of A, C, G and U nucleotides. As used herein, references to a polynucleotide comprising a T is considered to be a thymidine nucleotide in DNA and a uridine nucleotide in RNA, unless explicitly defined otherwise. Other naturally-occurring, non-naturally occurring, modified and / or synthetic nucleotides may also be present in the polynucleotides described herein, particularly nucleotides that may improve stability, transcription or translation of the polynucleotide described herein. For example, the polynucleotide may include inosine (I), 5’methylcytidine (5meC), N6-methyladenosine, pseudouridine, N1- methylpseudouridine, deoxyuridine (dU), abasic nucleotides, threose nucleotides (TNA), glycerol nucleotides (GNA), locked nucleotides (LNA) and peptide nucleotides (PNA). In some cases, then polynucleotide may comprise 5’methylcytidine (5meC), N6- methyladenosine, pseudouridine and / or N1-methylpseudouridine. Segmented poly(A) tail The invention provides a polynucleotide comprising a nucleic acid sequence encoding or comprising a segmented poly(A) tail. The segmented poly(A) tail may comprise at least 80 A nucleotides separated by at least two spacer sequences, wherein the spacer sequences are separated by a sequence of 11- 35 consecutive A nucleotides. In some cases, the segmented poly(A) tail consists of, or consists essentially of, at least 80 A nucleotides separated by at least two spacer sequences, wherein the spacer sequences are separated by a sequence of 11-35 consecutive A nucleotides. The segmented poly(A) tail typically comprises at least around 100 A nucleotides, such as at least around 150 A nucleotides, at least around 198 A nucleotides, at least around 200 A nucleotides, at least around 250 A nucleotides, or at least around 300 A nucleotides. The segmented poly(A) tail may comprise, for example, up to around 500 A nucleotides, such as up to around 400 A nucleotides, up to around 350 A nucleotides, up to around 300 A nucleotides, up to around 250 A nucleotides or up to around 200 A nucleotides. The segmented poly(A) tail may comprise 80-500 A nucleotides, such as 80-400 A nucleotides, 80-350 A nucleotides, 80-300 A nucleotides, 80-250 A nucleotides, 80-200 A nucleotides, 100-500 A nucleotides, 100-400 A nucleotides, 100- 350 A nucleotides, 100-300 A nucleotides, 100-250 A nucleotides or 100-200 A nucleotides. The segmented poly(A) tail may comprise an initial stretch of at least 11 consecutive A nucleotides (i.e.5’ of the first spacer in the segmented poly(A) tail), such as at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29 or at least 30 consecutive A nucleotides. The segmented poly(A) tail may comprise an initial stretch of up to 35 consecutive A nucleotides, such as up to 34, up to 33, up to 32, up to 31 or up to 30 consecutive A nucleotides. The segmented poly(A) tail may comprise an initial stretch of 11-35 consecutive A nucleotides, such as 22-35, 23-35, 25-35, 27-35, 29-35, 11-33, 22-33, 23-33, 25-33, 27-33, 29-33, 11-31, 22-31, 23-31, 25-31, 27-31, 29- 31 or around 30 consecutive A nucleotides. The segmented poly(A) tail may comprise a final stretch of at least 11 consecutive A nucleotides (i.e.3’ of the last spacer in the segmented poly(A) tail), such as at least 22, at least 23, at least 24, at least 25 or at least 26 consecutive A nucleotides. The segmented poly(A) tail may comprise a final stretch of up to 35 consecutive A nucleotides, such as up to 34, up to 33, up to 32, up to 31, up to 30, up to 29, or up to 28 consecutive A nucleotides. The segmented poly(A) tail may comprise a final stretch of 11-35 consecutive A nucleotides, such as 22-35, 23-35, 25-35, 26-35, 11-33, 22-33, 23-33, 25- 33, 26-33, 11-31, 22-31, 23-31, 25-31, 26-31, 11-30, 22-30, 23-30, 25-30, 26-30, 11-28, 22-28, 23-28, 25-28 or 26-28, or around 27 A nucleotides. The final stretch of consecutive A nucleotides may correspond to the complete binding footprint of PABPC1, i.e.22-35 consecutive A nucleotides as described herein, or a multiple thereof. Accordingly, the final stretch may be 22-35, 44-70, 66-105 or 88-120 consecutive A nucleotides. Alternatively, the final stretch may be 23-35, 46-70, 69-105 or 92-120 consecutive A nucleotides. In some cases, the final stretch may be around 27, around 54, around 81 or around 108 consecutive A nucleotides. In some cases, the final stretch may comprise at most 80 consecutive A nucleotides. In some cases, the segmented poly(A) tail may comprise no stretch of A nucleotides longer than 40 consecutive A nucleotides, such as no stretch of A nucleotides longer than 35 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 3 spacer sequences. For example, the segmented poly(A) tail may comprise at least 4 spacer sequences, at least 5 spacer sequences or at least 6 spacer sequences. The spacer sequences are separated by a sequence of 11-35 consecutive A nucleotides. For example, the spacer sequences may be separated by at least 22, at least 23, at least 24, at least 25, at least 26, or at least 27 consecutive A nucleotides. The spacer sequences may be separated by up to 35 consecutive A nucleotides, such as up to 34, up to 33, up to 32, up to 31, up to 30, up to 29, up to 28, or up to 27 consecutive A nucleotides. The spacer sequences may be separated by 22-35, 23-35, 25-35, 26-35, 11-33, 22-33, 23-33, 25-33, 26-33, 11-31, 22- 31, 23-31, 25-31, 26-31, 11-30, 22-30, 23-30, 25-30, 26-30, 11-28, 22-28, 23-28, 25-28 or 26-28 consecutive A nucleotides, or around 27 consecutive A nucleotides. For a segmented poly(A) tail comprising at least 3 spacer sequences, the spacer sequences may be separated by identical lengths of consecutive A nucleotides or different lengths of consecutive A nucleotides. The segmented poly(A) tail may comprise at least 3 or more spacer sequences, wherein at least two of the spacer sequences are separated by a sequence of 11-35 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 120 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22- 35 consecutive A nucleotides, and wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides. In some cases, the segmented poly(A) tail consists of, or consists essentially of, at least 120 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22-35 consecutive A nucleotides, and wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides. In some cases, the segmented poly(A) tail may comprise no stretch of A nucleotides longer than 35 consecutive A nucleotides. The segmented poly(A) tail typically comprises at least around 130 A nucleotides, such as at least around 140 A nucleotides, at least around 150 A nucleotides, at least around 160 A nucleotides, at least around 170 A nucleotides, at least around 180 A nucleotides, at least around 190 A nucleotides, at least around 198 A nucleotides, at least around 200 A nucleotides, at least around 250 A nucleotides, or at least around 300 A nucleotides. The segmented poly(A) tail may comprise, for example, up to around 500 A nucleotides, such as up to around 400 A nucleotides, up to around 350 A nucleotides, up to around 300 A nucleotides, up to around 260 A nucleotides, up to around 250 A nucleotides, up to around 240 A nucleotides, up to around 230 A nucleotides, up to around 22 A nucleotides, up to around 210 A nucleotides, or up to around 200 A nucleotides. The segmented poly(A) tail may comprise 120-500 A nucleotides, such as 120-400 A nucleotides, 120-350 A nucleotides, 120-300 A nucleotides, 120-260 A nucleotides, 120- 250 A nucleotides, 120-240 A nucleotides, 120-230 A nucleotides, 120-220 A nucleotides, 120-210 A nucleotides, 120-200 A nucleotides. The segmented poly(A) tail may comprise an initial stretch of at least 11 consecutive A nucleotides (i.e.5’ of the first spacer in the segmented poly(A) tail), such as at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29 or at least 30 consecutive A nucleotides. The segmented poly(A) tail may comprise an initial stretch of up to 35 consecutive A nucleotides, such as up to 34, up to 33, up to 32, up to 31 or up to 30 consecutive A nucleotides. The segmented poly(A) tail may comprise an initial stretch of 11-35 consecutive A nucleotides, such as 22-35, 23-35, 25-35, 27-35, 29-35, 11-33, 22-33, 23-33, 25-33, 27-33, 29-33, 11-31, 22-31, 23-31, 25-31, 27-31, 29- 31 or around 30 consecutive A nucleotides. The segmented poly(A) tail may comprise a final stretch of at least 11 consecutive A nucleotides (i.e.3’ of the last spacer in the segmented poly(A) tail), such as at least 22, at least 23, at least 24, at least 25 or at least 26 consecutive A nucleotides. The segmented poly(A) tail may comprise a final stretch of up to 35 consecutive A nucleotides, such as up to 34, up to 33, up to 32, up to 31, up to 30, up to 29, or up to 28 consecutive A nucleotides. The segmented poly(A) tail may comprise a final stretch of 11-35 consecutive A nucleotides, such as 22-35, 23-35, 25-35, 26-35, 11-33, 22-33, 23-33, 25- 33, 26-33, 11-31, 22-31, 23-31, 25-31, 26-31, 11-30, 22-30, 23-30, 25-30, 26-30, 11-28, 22-28, 23-28, 25-28 or 26-28, or around 27 A nucleotides. The final stretch of consecutive A nucleotides may correspond to the complete binding footprint of PABPC1, i.e.22-35 consecutive A nucleotides as described herein. The segmented poly(A) tail may comprise at least 4 spacer sequences. For example, the segmented poly(A) tail may comprise at least 5 spacer sequences or at least 6 spacer sequences. The spacer sequences may be separated by at least 23, at least 24, at least 25, at least 26, or at least 27 consecutive A nucleotides. The spacer sequences may be separated by up to 34, up to 33, up to 32, up to 31, up to 30, up to 29, up to 28, or up to 27 consecutive A nucleotides. The spacer sequences may be separated by 23-35, 25-35, 26-35, 11-33, 22-33, 23-33, 25-33, 26-33, 11-31, 22-31, 23-31, 25-31, 26-31, 11-30, 22- 30, 23-30, 25-30, 26-30, 11-28, 22-28, 23-28, 25-28 or 26-28 consecutive A nucleotides, or around 27 consecutive A nucleotides. The spacer sequences may be separated by identical lengths of consecutive A nucleotides or different lengths of consecutive A nucleotides. The segmented poly(A) tail may comprise at least 120 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22- 30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 120 A nucleotides separated by at least four spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 120 A nucleotides separated by at least five spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 120 A nucleotides separated by at least six spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23- 30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 130 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22- 30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 130 A nucleotides separated by at least four spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 130 A nucleotides separated by at least five spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 130 A nucleotides separated by at least six spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23- 30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 140 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22- 30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 140 A nucleotides separated by at least four spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 140 A nucleotides separated by at least five spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 140 A nucleotides separated by at least six spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23- 30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 150 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22- 30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 150 A nucleotides separated by at least four spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 150 A nucleotides separated by at least five spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 150 A nucleotides separated by at least six spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23- 30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 160 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22- 30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 160 A nucleotides separated by at least four spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 160 A nucleotides separated by at least five spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 160 A nucleotides separated by at least six spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23- 30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 170 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22- 30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 170 A nucleotides separated by at least four spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 170 A nucleotides separated by at least five spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 170 A nucleotides separated by at least six spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23- 30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 180 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22- 30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 180 A nucleotides separated by at least four spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 180 A nucleotides separated by at least five spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 180 A nucleotides separated by at least six spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23- 30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 190 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22- 30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 190 A nucleotides separated by at least four spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 190 A nucleotides separated by at least five spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides. The segmented poly(A) tail may comprise at least 190 A nucleotides separated by at least six spacer sequences, wherein the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as 23- 30, 25-29, 26-28 or around 27 consecutive A nucleotides. In some cases, the segmented poly(A) tail comprises a sequence of the formula AX(SAY)z, wherein S is a spacer sequence, X is 11-35, Y is 11-35 and Z is at least 2. In other words, the segmented poly(A) tail comprises a sequence of the formula A11-35(SA11-35)at least 2. In some cases, the segmented poly(A) tail consists of, or consists essentially of a sequence of the formula AX(SAY)z, wherein S is a spacer sequence, X is 11-35, Y is 11- 35 and Z is at least 2. For example, for a segmented poly(A) tail comprising a sequence of the formula A30(SA27)3, wherein S is a single G nucleotide, the sequence of the segmented poly(A) tail comprises AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAGAAAAAAAAAAAAAAAAAA AAAAAAAAAGAAAAAAAAAAAAAAAAAAAAAAAAAAAGAAAAAAAAAAA AAAAAAAAAAAAAAAA X is at least 11. For example, X may be at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29 or at least 30. X is up to 35. For example, X, may be up to 34, up to 33, up to 32, up to 31 or up to 30. X may be 22-35, 23-35, 25-35, 27-35, 29-35, 11-33, 22-33, 23-33, 25-33, 27-33, 29-33, 11-31, 22-31, 23-31, 25-31, 27- 31, 29-31 or around 30. X therefore defines the length of an initial stretch of consecutive A nucleotides in the segmented poly(A) tail. However, it is possible that a further stretch of consecutive A nucleotides precedes the segmented poly(A) tail as defined herein. Y is at least 11. For example, Y may be at least 22, at least 23, at least 24, at least 25, at least 26, or at least 27. Y is up to 35. For example, Y may be up to 34, up to 33, up to 32, up to 31, up to 30, up to 29, up to 28, or up to 27. Y may be 22-35, 23-35, 25-35, 26- 35, 11-33, 22-33, 23-33, 25-33, 26-33, 11-31, 22-31, 23-31, 25-31, 26-31, 11-30, 22-30, 23-30, 25-30, 26-30, 11-28, 22-28, 23-28, 25-28 or 26-28, or around 27. Z is at least 2. For example, Z may be at least 3, at least 4, at least 5 or at least 6. In some cases, the segmented poly(A) tail may comprise a further nucleic acid sequence 3’ to the formula AX(SAY)z. For example, the segmented poly(A) tail may comprise further spacer sequences and / or consecutive stretches of A nucleotides. Accordingly, the segmented poly(A) tail may comprise a sequence of the formula AX(SAY)zAN. In some cases, the segmented poly(A) tail consists of, or consists essentially of a sequence of the formula AX(SAY)zAN, wherein S is a spacer sequence, X is 11-35, Y is 11-35 and Z is at least 2. N may be at least 10. Typically, N corresponds to the complete binding footprint of PABPC1, i.e.22-35 consecutive A nucleotides as described herein, or a multiple thereof. Accordingly, N may be 22-35, 44-70, 66-105 or 88-120. Alternatively, N may be 23-35, 46-70, 69-105 or 92-120. In some cases, N is around 27, around 54, around 81 or around 108. In some cases, N is at most 80. In some cases, the segmented poly(A) tail comprises a sequence of the formula AX(SAY)z, wherein S is a spacer sequence, X is 22-35, Y is 22-35 and Z is at least 4, and wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides. In other words, the segmented poly(A) tail comprises a sequence of the formula A22-35(SA22-35)at least 4,wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides. In some cases, the segmented poly(A) tail consists of, or consists essentially of a sequence of the formula AX(SAY)z, wherein S is a spacer sequence, X is 22-35, Y is 22-35 and Z is at least 4. X may be at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29 or at least 30. X is up to 35. For example, X, may be up to 34, up to 33, up to 32, up to 31 or up to 30. X may be 23-35, 25-35, 27-35, 29-35, 11-33, 22-33, 23-33, 25-33, 27-33, 29-33, 11-31, 22-31, 23-31, 25-31, 27-31, 29-31 or around 30. Y may be at least 23, at least 24, at least 25, at least 26, or at least 27. Y is up to 35. For example, Y may be up to 34, up to 33, up to 32, up to 31, up to 30, up to 29, up to 28, or up to 27. Y may be 23-35, 25-35, 26-35, 11-33, 22-33, 23-33, 25-33, 26-33, 11-31, 22- 31, 23-31, 25-31, 26-31, 11-30, 22-30, 23-30, 25-30, 26-30, 11-28, 22-28, 23-28, 25-28 or 26-28, or around 27. Z may be at least 5 or at least 6. The segmented poly(A) tail may comprise a sequence of the formula A22-35(SA22-35)at least4, A22-35(SA22-35)at least 5, A22-35(SA22-35)at least 6, A22-35(SA22-35)at least 7, A22-35(SA22-35)at least 8, A23-35(SA23-35)at least 4, A23-35(SA23-35)at least 5, A23-35(SA23-35)at least 6, A23-35(SA23-35)at least 7, A23-35(SA23-35)at least 8, A25-35(SA25-35)at least 4, A25-35(SA25-35)at least 5, A25-35(SA25-35)at least 6, A25-35(SA25-35)at least 7, A25-35(SA25-35)at least 8, A23-30(SA23-30)at least 4, A23-30(SA23-30)at least 5, A23-30(SA23-30)at least 6, A23-30(SA23-30)at least 7, A23-30(SA23-30)at least 8, A25-30(SA25-30)at least 4, A25-30(SA25-30)at least 5, A25-30(SA25-30)at least 6, A25-30(SA25-30)at least 7, A25-30(SA25-30)at least 8, A28-32(SA25-29)at least 4, A28-32(SA25-29)at least 5, A28-32(SA25-29)at least 6, A28-32(SA25-29)at least 7, A28-32(SA25-29)at least 8, A29-31(SA26-28)at least 4, A29-31(SA26-28)at least 5, A29-31(SA26-28)at least 6, A29-31(SA26-28)at least 7, A29-31(SA26-28)at least 8, A30(SA27)at least 4, A30(SA27)at least 5, A30(SA27)atleast 6, A30(SA27)at least 7, or A30(SA27)at least 8, wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides. The segmented poly(A) tail may consist of a sequence of the formula A22-35(SA22-35)at least 4, A22-35(SA22-35)at least 5, A22-35(SA22-35)at least 6, A22-35(SA22-35)at least 7, A22-35(SA22-35)at least 8, A23-35(SA23-35)at least 4, A23-35(SA23-35)at least 5, A23-35(SA23-35)at least 6, A23-35(SA23-35)at least 7, A23-35(SA23-35)at least 8, A25-35(SA25-35)at least 4, A25-35(SA25-35)at least 5, A25-35(SA25-35)at least 6, A25-35(SA25-35)at least 7, A25-35(SA25-35)at least 8, A23-30(SA23-30)at least 4, A23-30(SA23-30)at least 5, A23-30(SA23-30)at least 6, A23-30(SA23-30)at least 7, A23-30(SA23-30)at least 8, A25-30(SA25-30)at least 4, A25-30(SA25-30)at least 5, A25-30(SA25-30)at least 6, A25-30(SA25-30)at least 7, A25-30(SA25-30)at least 8, A28-32(SA25-29)at least 4, A28-32(SA25-29)at least 5, A28-32(SA25-29)at least 6, A28-32(SA25-29)at least 7, A28-32(SA25-29)at least 8, A29-31(SA26-28)at least 4, A29-31(SA26-28)at least 5, A29-31(SA26-28)at least 6, A29-31(SA26-28)at least 7, A29-31(SA26-28)at least 8, A30(SA27)at least 4, A30(SA27)at least 5, A30(SA27)at least 6, A30(SA27)at least 7, or A30(SA27)at least 8. The segmented poly(A) tail is typically considered to start at the first sequence of, or encoding, at least 11 consecutive A nucleotides in the nucleic acid sequence, such as at least 22, at least 23, at least 24, at least 25, at least 26 or at least 37 consecutive A nucleotides in the nucleic acid sequence. The segmented poly(A) tail is typically considered to end at the last sequence of, or encoding, at least 11 consecutive A nucleotides in the nucleic acid sequence, such as at least 22, at least 23, at least 24, at least 25, at least 26 or at least 37 consecutive A nucleotides in the nucleic acid sequence. Without being bound by theory, sequences of consecutive A nucleotides smaller than the 11 consecutive A nucleotides have severely reduced binding to PABPC1 and therefore reduced functionality, whereas sequences of 22 or more consecutive A nucleotides, approximately corresponding to the full binding footprint of PABPC1, have favourable binding to PABPC1. In some cases, the nucleic acid sequence may comprise, or encode, an additional sequence after the segmented poly(A) tail, for example, 1-10 nucleotides. The additional sequence, may, for example, protect the segmented poly(A) tail from deadenylation. Spacer sequences The segmented poly(A) tail comprises at least two spacer sequences. Each spacer sequence may comprise an identical nucleic acid sequence to each other spacer sequence in the segmented poly(A) tail. Two or more spacers sequences in the segmented poly(A) tail may comprise different nucleic acid sequences. For example, each of the spacer sequences in the segmented poly(A) tail may comprise a different nucleic acid sequence. A spacer sequence is typically 1-50 nucleotides in length, i.e. consecutive nucleotides, preferably 1-25 nucleotides in length. In some cases, the spacer sequence is 2-25 nucleotides in length, i.e. consecutive nucleotides. Without being bound by theory, it is understood that if the spacer sequence is too long, the gaps between sites suitable for binding by PABPC1 increase and consequently reduce the poly(A) tail’s functionality. In some cases, the spacer sequence is up to 50 nucleotides in length, such as up to 30, up to 25, up to 20, up to 15, up to 12, up to 10, up to 8, up to 6, up to 5, up to 4, up to 3 or up to 2 nucleotides in length. The spacer sequence may be at least 3 nucleotides in length, such as at least 4, at least 5, at least 6, at least 7, at least 10 or at least 15 nucleotides in length. Preferably, the spacer sequence is a single nucleotide. Without being bound by theory, the purpose of the spacer sequence is to reduce the likelihood of recombination during the amplification and / or subcloning of a polynucleotide encoding the segmented poly(A) tail and / or to reduce the susceptibility of the segmented poly(A) tail to deadenylation enzymes (Lim et al., 2018). It is preferred that the number of A nucleotides in the spacer is minimised. In some cases, the spacer sequence does not comprise an A nucleotide at the first or last position of the spacer, i.e. an A nucleotide is not present at the 5’ or 3’ end of the spacer sequence. The spacer sequence may comprise a B nucleotide (i.e. C, G or T) at the first and last position of the spacer. The spacer sequence may comprise a K nucleotide (i.e. G or T) at the first and last position of the spacer. The spacer sequence may comprise a G nucleotide at the first and last position of the spacer. In some cases, the spacer sequence does not comprise three or more, or two or more consecutive A nucleotides. Preferably, the spacer sequence does not comprise an A nucleotide. For example, the spacer sequence may consist of B nucleotides (i.e. C, G or T), such as a single B nucleotide. More preferably, the spacer sequence may consist of K nucleotides (i.e. G or T), such as a single K nucleotide. Most preferably, the spacer sequence may consist of G nucleotides, such as a single G nucleotide. Where the segmented poly(A) tail comprises a stretch of at least 11, but less than 22, consecutive A nucleotides, a spacer sequence adjacent to said stretch preferably consists of a single nucleotide, such as a single G. Without being bound by theory, a single PABPC1 molecule may tolerate binding to a sequence comprising a spacer sequence where the pairs of RRM domains bind to a stretch of at least 11 consecutive A nucleotides on either side of the spacer. Where the spacer sequence is two or more nucleotides in length, the spacer sequence may be TGCAT, GATATC, a poly(C) sequence (such as C30) or a histone-stem loop sequence (as described in WO 2016 / 091391 A1). In some cases, the spacer sequence may be a sequence of around 10 consecutive random nucleotides, typically comprising an equal distribution of A, G, C and T (as described in WO 2015 / 005324). Accordingly, in some cases, the segmented poly(A) tail may comprise one of the sequences exemplified in Figure 2: A30GA27GA27A30GA27GA27GA27A30GA27GA27GA27GA27A30GA27GA27GA27GA27GA27A30GA27GA27GA27GA27GA27GA27In some cases, one or more of the G spacer sequences in the segmented poly(A) tail sequences provided above, may be replaced with a different spacer sequence, such as TGCAT, GATATC, a poly(C) sequence, a histone-stem loop sequence or a sequence of around 10 consecutive random nucleotides. Other features of the nucleic acid sequence The polynucleotide comprises a nucleic acid sequence encoding or comprising a segmented poly(A) tail. The segmented poly(A) tail is typically at the 3’ end of the sequence of, or encoded by, the nucleic acid sequence. This is to mean that when the nucleic acid sequence is transcribed, for example by a DNA-dependent RNA polymerase or an RNA-dependent RNA polymerase, the transcribed sequence comprises the segmented poly(A) tail at its 3’ end. Typically, the sequence of, or encoded by, the nucleic acid molecule does not comprise any sequence elements 3’ of the segmented poly(A) tail. However, in some cases, the sequence of, or encoded by, the nucleic acid molecule may comprise a sequence 3’ of the segmented poly(A) tail. The sequence 3’ of the segmented poly(A) tail may be non-functional. The sequence typically does not interfere or reduce the function of the segmented poly(A) tail. The sequence, may, for example, be an overhang or blunt-ended tail deriving from restriction endonuclease digestion of the polynucleotide prior to transcription. The sequence may be up to 20 nucleotides in length, such as up to 10, up to 9, up to 8, up to 7, up to 6, up to 5, up to 4, up to 3, up to 2, or 1 nucleotide in length. The nucleic acid sequence may encode or comprise further elements. For example, the sequence of, or encoded by, the nucleic acid sequence may comprise a 5’ untranslated region (5’ UTR), a 3’ untranslated region (3’ UTR) and / or an open reading frame (ORF), a sequence for introducing an ORF, or a sequence encoding a peptide, polypeptide or protein. The term “peptide” typically refers to a molecule comprising 2 or more consecutive amino acids linked by peptide bonds, more typically 2 to 50 consecutive amino acids. The term “polypeptide” typically refers to a molecule comprising 50 or more consecutive amino acids linked by peptide bonds. There is no practical upper limit to the length of the polypeptide. The term “protein” can typically be used interchangeably with the term “polypeptide”, but also encompass structures formed of more than one polypeptide chain, such as via cleavage and / or disulphide bond formation. The peptide, polypeptide or protein may comprise substances that are not amino acids, such as post-translational modifications, covalently- or non-covalently-bound co-factors and the like. The 5’ UTR, the 3’ UTR and / or the ORF, a sequence for introducing an ORF, or a sequence encoding a peptide, polypeptide or protein, are typically present in the sequence of, or encoded by, the nucleic acid sequence 5’ of the segmented poly(A) tail. For example, the sequence of, or encoded by, the nucleic acid sequence may comprise, in order, the 5’ UTR, the ORF, the sequence for introducing an ORF, or the sequence encoding a peptide, polypeptide or protein, the 3’ UTR and the segmented poly(A) tail. In this context, the term “in order” refers to the arrangement of features in the resulting sequence. If the nucleic acid sequence is present on an antisense strand of the polynucleotide encoding the features, then the nucleic acid sequence may comprise in the 3’ to 5’ direction a sequence encoding the 5’ UTR, a sequence encoding the ORF, a sequence encoding the 3’ UTR and a sequence encoding the segmented poly(A) tail. An RNA molecule encoded by the nucleic acid sequence, or the RNA polynucleotide comprising the nucleic acid sequence, may comprise in a 5’ to 3’ direction the 5’ UTR, the ORF, the 3’ UTR and the segmented poly(A) tail. The 5’ UTR is typically the nucleic acid sequence between the transcription start site and the ORF. The 5’ UTR may comprise a ribosomal binding site, such as the Kozak sequence or translation initiation site. The 5’ UTR may comprise an internal ribosome entry site (IRES), as described further herein. The 5’ UTR may comprise other regulatory elements such as one or more upstream ORFs (uORFs) and / or secondary structure elements, such as hairpin-loops. In some cases, the 5’UTR does not comprise a uORF. A uORF is typically a nucleic acid sequence comprising an AUG start codon, and optionally comprising a stop codon prior to the start codon of the main ORF. The presence of a uORF in the 5’UTR may attenuate translation of the subsequent ORF. The 5’ UTR may comprise a 5’UTR or fragment thereof of a TOP gene, such as a 5’ terminal oligopyrimidine tract (TOP) gene encoding a ribosomal large protein, optionally lacking the 5’ TOP motif. The ORF typically encodes an amino acid sequence. The ORF may also encode non- protein coding sequences, such as introns or other regulatory elements. The ORF typically begins with a start codon and terminates with a stop codon. The ORF may encode a peptide, polypeptide or protein. The ORF may encode a therapeutic peptide, polypeptide or protein. Therapeutic peptides, polypeptides and proteins are well-known to the skilled person, and may be useful for the prevention or treatment of an inherited or acquired disease. The ORF may encode an antigen. An antigen may be considered a therapeutic peptide, polypeptide or protein that achieves the therapeutic effect by eliciting an immune response in a host, such as a human host. The antigen may be a viral, bacterial, fungal, parasitic, allergenic, autoimmune or tumorigenic antigen. The viral, bacterial, fungal or parasitic antigen may be derived from a virus, bacteria, fungus or parasite that is a pathogen, e.g. a human pathogen. For example, the viral, bacterial, fungal or parasitic antigen may be derived from an infectious virus, bacteria, fungus or parasite, e.g. a virus, bacteria, fungus or parasite that is capable of infecting humans. The sequence for introducing an ORF may be a multiple cloning site. The multiple cloning site typically comprises one or more restriction endonuclease sites that are unique to the polynucleotide, typically two or more, three or more, four or more or five or more restriction endonuclease sites that are unique to the polynucleotide. The 3’ UTR is typically the nucleic acid sequence between the ORF and the segmented poly(A) tail. The 3’ UTR may comprise regulatory elements such as one or more inverted repeats that are capable of forming a stem-loop structure. Stem-loop structures are known to act as a barrier for exoribonucleases and / or interact with proteins known to increase RNA stability. The 3’ UTR may comprise a binding site for a regulatory protein and / or an miRNA. The 3’ UTR may comprise an AU-rich element (ARE). The 3’ UTR typically functions to enhance protein expression and / or to improve the half-life of the transcribed polynucleotide, for example, when compared to a control lacking a 3’ UTR. The 3’ UTR may comprise the 3’UTR of an albumin gene, an α-globin gene, a β-globin gene, a ribosomal protein gene, a tyrosine hydroxylase gene, a lipoxygenase gene, or a collagen alpha gene. The sequence of, or encoded by, the nucleic acid sequence may comprise a non-coding functional RNA sequence. The non-coding functional RNA sequence is typically present in the sequence of, or encoded by, the nucleic acid sequence 5’ of the segmented poly(A) tail. For example, the sequence of, or encoded by, the nucleic acid may comprise, in order, the non-coding functional RNA sequence and the segmented poly(A) tail. The sequence of, or encoded by, the nucleic acid sequence may comprise a non-coding functional RNA sequence, a 5’UTR, a 3’UTR and / or an ORF, a sequence for introducing an ORF, or a sequence encoding a peptide, polypeptide or protein. In some cases, the non-coding functional RNA sequence is present within the 5’ UTR and / or the 3’UTR. In some cases, sequence of, or encoded by, the nucleic acid may comprise the non-coding functional RNA sequence prior to or after a 5’ UTR, prior to or after an ORF, a sequence for introducing an ORF, or a sequence encoding a peptide, polypeptide or protein, or prior to or after a 3’ UTR. For example, the sequence of, or encoded by, the nucleic acid sequence may comprise, in order, the 5’ UTR, the non-coding functional RNA sequence, the 3’ UTR and the segmented poly(A) tail. The non-coding functional RNA sequence may comprise an RNA sequence forming part of a ribonucleoprotein, a ribozyme, ribosomal RNA, transfer RNA, small nuclear RNA, small nucleolar RNA, Y RNA, microRNA, antisense RNA, and / or an RNA sequence that is capable of sequestering miRNAs or ribosomal binding proteins. The antisense RNA may be capable of hybridising to a target sense transcript, to thereby inhibit further transcription or translation of said sense transcript. The RNA sequence that is capable of sequestering miRNAs and / or ribosomal binding proteins, may comprise a sequence capable of hybridising to miRNAs and / or a sequence that acts as a binding site for ribosomal binding proteins. Without being limited to theory, it is believed that such non- coding functional RNA sequences may control expression of other RNA molecules. The sequence of, or encoded by, the nucleic acid sequence may comprise an internal ribosome entry site (IRES). For example, the sequence of, or encoded by, the nucleic acid sequence may comprise an IRES, and an ORF, a sequence for introducing an ORF, or a sequence encoding a peptide, polypeptide or protein. The sequence of, or encoded by, the nucleic acid sequence may comprise an IRES, a 5’ UTR, a 3’ UTR, and / or an ORF, a sequence for introducing an ORF, or a sequence encoding a peptide, polypeptide or protein. The IRES is typically present in the sequence of, or encoded by, the nucleic acid sequence 5’ of the segmented poly(A) tail. The sequence of, or encoded by, the nucleic acid sequence may comprise, in order, the IRES, the ORF, the sequence for introducing an ORF, or the sequence encoding a peptide, polypeptide or protein, and the segmented poly(A) tail. The sequence of, or encoded by, the nucleic acid sequence may comprise two ORFs, two sequences for introducing an ORF or two sequences encoding a peptide, polypeptide or protein, and an IRES located between the two ORFs, two sequences for introducing an ORF or two sequences encoding a peptide, polypeptide or protein. In some cases, the IRES is present in the 5’ UTR, as described herein. Accordingly, the sequence of, or encoded by, the nucleic acid sequence may comprise, in order, a 5’ UTR comprising the IRES, the ORF, the sequence for introducing an ORF, or the sequence encoding a peptide, polypeptide or protein, and the segmented poly(A) tail. The IRES may, for example, be derived from an IRES of a viral genome or from a eukaryotic genome. Examples of IRESs derived from a viral genome include the picornavirus IRES, apthovirus IRES, Kaposi’s sarcoma-associated herpesvirus IRES, hepatitis A IRES, hepatitis C IRES, pestivirus IRES, Cripavirus RES, Rhopalosiphum padi virus IRES or MDV IRES. Examples of IRESs derived from a eukaryotic genome include IRESs found in the mRNA of FGF-1, FGF-2, PDGF, VEGF, IGF-II, Antennapedia, Ultrabithorax, MYT-2, NF-κB repressing factor NRF, AML1 / RUNX1, Gtx homeodomain protein, eIF4Ga eIF4Gia, eIF4G2, c-myc, L-myc, Pim-1, Protein kinase p58PITSLRE, p53, SLC7A1,Cat-1, Notch 2, Voltage-gated potassium channel, Apaf-1, XIAP, HIAP2, Bcl-xL, Bcl-2, ARC, α-subunit of calcium calmodulin dependent kinase II dendrin, MAP2, RC3, Amyloid precursor protein, BiP, HSP70, β-subunit of mitochondrial H+-ATP synthase, Ornithine decarboxylase, connexins 32 and 43, HIF-1α and APC. The nucleic acid sequence is typically transcribed to produce a common transcript. It is, however, envisaged, that a single nucleic acid sequence may give rise to two or more difference transcribed molecules, for example, due to co- and / or post-transcription processing, such as alternative splicing. Other features of the polynucleotide The polynucleotide comprising a nucleic acid sequence encoding a segmented poly(A) tail typically comprises one or more further sequence elements. The polynucleotide may further comprise a promoter. The promoter aids in the transcription of the nucleic acid sequence. The promoter may be the T7 promoter, the SP6 promoter or the T3 promoter. The promoter is typically upstream of the nucleic acid sequence on the polynucleotide. For example, the promoter is 5’ of the nucleic acid sequence on a sense strand, and 3’ of the nucleic acid sequence on an antisense strand. The polynucleotide may further comprise a marker gene. The marker gene typically encodes a polynucleotide or amino acid sequence that enables a cell comprising the polynucleotide to be distinguished from a cell that does not comprise the polynucleotide. The marker gene may be a screenable marker gene or a selectable marker gene. The screenable marker gene may encode a fluorescent protein such as green fluorescent protein or a variant thereof. The screenable marker gene may be a bacterial LacZ gene. The selectable marker gene may be an antibiotic resistance gene, such as a gene encoding β-lactamase. The polynucleotide may further comprise an origin of replication (ori). The original of replication may be a single origin of chromosomal replication (oriC). The oriC is typically a bacterial oriC, such as E. coli oriC. The ori may be ColE1, pMB1, a derivative of PMB1 such as that found in the pUC vector, pBR322, pSC101, R6K, p15A or F1 ori. The polynucleotide may further comprise a multiple cloning site (MCS). An MCS typically comprises two or more unique restriction endonuclease recognition sites for allowing a nucleic acid sequence to be inserted into the polynucleotide. The polynucleotide preferably comprises a restriction endonuclease recognition site (restriction site), typically at the terminal end of the segmented poly(A) tail encoded by the nucleic acid sequence. In other words, cleavage at the restriction site does not separate the segmented poly(A) tail encoded by the nucleic acid sequence from other sequence elements, as described herein, encoded by the nucleic acid sequence. For example, if the polynucleotide is a double-stranded DNA polynucleotide, the restriction site is typically at the 3’ end of the sequence encoding the segmented poly(A) tail on the sense strand and at the 5’ end of the sequence encoding the segmented poly(A) tail on the antisense strand. The purpose of the restriction site is typically to linearise the polynucleotide for in vitro transcription. The restriction site may partially or completely overlap the sequence encoding the segmented poly(A) tail. In some cases, the restriction site is a SapI restriction site: 5’…G C T C T T C (N)1˅ … 3’ 3’ …C G A G A A G (N)4^ … 5’ The polynucleotide may therefore be designed such that cleavage at the restriction site does not leave any nucleotides that do not encode the segmented poly(A) tail at the terminal end of the linear polynucleotide, or minimises the number of nucleotides that do not encode the segmented poly(A) tail at the terminal end of the linear polynucleotide. In some cases, the restriction site is located -5 to 50 nucleotides, such as 0-50, 1-50, 0-26, 5- 26 or 24-26 nucleotides from the terminal end of the segmented poly(A) tail encoded by the nucleic acid sequence. The restriction site is preferably a sequence that is capable of being recognised and cleaved by a restriction endonuclease that results in a 5’ overhang or a blunt end. In some cases, the restriction site is a sequence that is capable of being recognised and cleaved by the restriction endonuclease SapI, StuI, SspI or XbaI, or isoschizomers thereof. The polynucleotide is suitable, typically after linearization, for in vitro transcription of RNA, such as of mRNA. In some cases, the polynucleotide comprises no stretch of A nucleotides longer than 40 consecutive A nucleotides, such as no longer than 35 consecutive A nucleotides. In other words, the polynucleotide does not comprise a stretch of more than 40 consecutive A nucleotides, or does not comprise a stretch of more than 35 consecutive A nucleotides. Vectors The invention also provides a vector comprising a polynucleotide of the invention. The vector may be a linear vector or a circular vector. The vector is typically a circular vector capable of being linearised by the action of a restriction endonuclease enzyme at a restriction site at the terminal end of the sequence encoding the segmented poly(A) tail, as described herein. The vector is typically suitable for in vitro transcription (IVT), i.e. to produce an RNA molecule encoded by the nucleic acid sequence. Cells The invention also provides a cell comprising a polynucleotide of the invention or a vector of the invention. Where the polynucleotide comprises a nucleic acid sequence encoding the segmented poly(A) tail, the cell is typically suitable for propagating the polynucleotide. For example, where the vector is a double-stranded circular DNA molecule comprising the polynucleotide, the cell may be suitable for producing more copies of the double- stranded circular DNA molecule. The cell may be a prokaryotic cell. The prokaryotic cell may be a bacterial cell. The bacterial cell may be an E. coli cell, such as an E. coli DH5alpha cell. The cell may comprise a mutation that reduces recombination. For example, the cell may be an E. coli cell comprising a mutation in the recA, recA1 and / or recA13 gene, such as deletion of the recA, recA1 and / or recA13 gene, and / or a mutation in the recBCD gene to reduce or abolish exonuclease V activity. Where the polynucleotide is an RNA molecule comprising a nucleic acid sequence comprising a segmented poly(A) tail, the cell may be suitable for transcribing a non- coding functional RNA sequence and / or translating a peptide, polypeptide or protein encoded by the nucleic acid molecule. For example, where the RNA molecule encodes a therapeutic peptide, polypeptide or protein, the cell may be suitable for translating the RNA molecule to produce the peptide, polypeptide or protein. The cell may be a eukaryotic cell. The eukaryotic cell may be a mammalian cell. The cell may be a human cell. The subject may be a non-human mammalian cell, such as a mouse, rat, cat, dog, pig, goat, sheep, horse, cow, camel or non-human primate cell. The cell may be isolated from a subject. The cell may be an immortal cell line. The cell may be an immune cell, such as professional antigen presenting cell (for example a dendritic cell, a monocyte or a macrophage). RNA molecules In some cases, the invention relates to a polynucleotide comprising a nucleic acid sequence comprising a segmented poly(A) tail as described herein. The polynucleotide may be a polyribonucleotide, also referred to herein as an RNA polynucleotide or RNA molecule. The RNA molecule may be obtained or obtainable by transcription of the nucleic acid sequence encoding a segmented poly(A) tail as described herein, or encoded by a nucleic acid sequences as described herein. The RNA molecule may be obtained by means other than transcription of the nucleic acid sequence as described herein. For example, the RNA molecule may be synthesised chemically. The RNA molecule comprises a segmented poly(A) tail. The segmented poly(A) tail may comprise at least 80 A nucleotides separated by at least two spacer sequences, wherein the spacer sequences are separated by a sequence of 19-35 consecutive A nucleotides. The segmented poly(A) tail may comprise a sequence of the formula AX(SAY)z, wherein S is a spacer sequence, X is 19-35, Y is 19-35 and Z is at least 2. The segmented poly(A) tail may be further defined as described herein in relation to the polynucleotide of the invention. The RNA molecule may be further defined as described herein in relation to the sequence of, or encoded by, the nucleic acid of the polynucleotide of the invention. For example, the RNA molecule may comprise, in the 5’ to 3’ direction, a 5’ UTR, an ORF, a 3’ UTR and the segmented poly(A) tail. The RNA molecule is typically an mRNA molecule. An mRNA molecule typically comprises, in the 5’ to 3’ direction, a 5’ UTR, an ORF, a 3’ UTR and the segmented poly(A) tail. The RNA molecule may comprise a 5’ cap. The 5’ cap may be suitable for binding to eukaryotic translation initiation factor 4E (eIF4E). The 5’ cap may be a 7- methylguanosine cap. In some cases, the 5’ cap may be a functional analogue of a 7- methylguanosine cap. The term “functional analogue” is intended to refer to a molecule at the 5’ end of the RNA molecule that retains the function of the 7-methylguanosine cap, e.g. to bind eIF4E and enable circularisation of the mRNA molecule. In some cases, the functional analogue may enhance the properties of the mRNA molecule when compared to the 7-methylguanosine cap, e.g. by enhancing the stability of the RNA molecule and / or recruitment of ribosomes. In some cases, the RNA molecule comprises a structure capable of initiating cap independent translation initiation. In some cases, the RNA molecule comprises the structure in addition to a 5’ cap. In some cases, the RNA molecule does not comprise a 5’ cap. The structure may be an IRES. The structure may be a cap independent translation enhancer (CITE, otherwise known as a cap independent translation element). A CITE typically binds mRNA-recruiting translational components, such as translation initiation factors and / or a 60S ribosomal subunit. The CITE may be comprised within the 5’ UTR and / or the 3’ UTR. The CITE may be a CITE comprised within a 3’UTR of an RNA virus, such as an RNA plant virus. The structure may be the presence of a N6- methyladenosine at the 5’ end of the RNA molecule and / or double stranded RNA regions at the 5’ end or 5’ UTR of the RNA molecule. In some cases, the RNA molecule comprises no stretch of A nucleotides longer than 40 consecutive A nucleotides, such as no longer than 35 consecutive A nucleotides. In other words, the RNA molecule does not comprise a stretch of more than 40 consecutive A nucleotides, or does not comprise a stretch of more than 35 consecutive A nucleotides. Pharmaceutical compositions The invention further provides a pharmaceutical composition comprising the RNA molecule of the invention. The pharmaceutical composition may further comprise a pharmaceutically acceptable excipient. For the RNA molecule to have pharmaceutical use, the RNA molecule typically encodes a therapeutic RNA sequence, a therapeutic peptide, a therapeutic polypeptide, a therapeutic protein or an antigen, as described herein. The pharmaceutical composition may thus be administered to a subject and the subject’s cells transcribe and / or translate the RNA molecule to produce a therapeutic molecule in vivo. The pharmaceutical composition may comprise further components to aid uptake of the RNA molecule by a subject’s cells, e.g. by transfection. The component typically sequesters or encapsulates the RNA molecule. For example, the pharmaceutical composition may comprise a micelle, such as a lipid or polymer micelle. The pharmaceutical composition may comprise a liposome or a lipoplex. The pharmaceutical composition may comprise a viral vector, such as an adeno-associated virus (AAV) vector. The pharmaceutical composition may comprise a polymeric nanoparticle. The pharmaceutical composition may comprise a lipid nanoparticle. The RNA molecule is typically encapsulated by the lipid nanoparticle. The suitable lipid nanoparticles are described, for example, in Hou et al., 2021. Currently approved lipid nanoparticles typically comprise a cationic or ionisable lipid, cholesterol, a helper lipid and a PEG- lipid. The pharmaceutical composition may comprise a polymer or polymer-based nanoparticle, such as poly(beta-amino ester)s. The pharmaceutical composition may comprise an exosome, suitable for delivering mRNA into a cell. The exosome is typically an exosome derived from a cell of the same genus or species as the subject. For example, the exosome may be a human embryonic kidney (HEK) cell-derived exosome, a bone marrow stem cells (BMSC)-derived exosome, a milk-derived exosome or a red- blood cell derived exosome. Further transfection agents are described in WO 2016 / 091391 A1. The RNA molecule is typically included in the pharmaceutical composition in an effective amount. The term “effective amount” is intended to refer to a quantity sufficient to achieve a measurable therapeutic response in a subject to which the pharmaceutical composition has been administered. Medical uses The invention also relates to the therapeutic use of the polynucleotide, vector, RNA molecule or pharmaceutical composition of the invention. The invention provides a method of treating or preventing a disease, disorder or condition in a subject, the method comprising administering the polynucleotide, vector, RNA molecule or pharmaceutical composition of the invention to the subject. The invention also provides the polynucleotide, vector, RNA molecule or pharmaceutical composition of the invention for use in a method of treating or preventing disease in a subject. The invention also relates to the use of the polynucleotide, vector, RNA molecule or pharmaceutical composition of the invention in a method of treating or preventing a disease in a subject, or use of the polynucleotide, vector, RNA molecule or pharmaceutical composition of the invention for treating or preventing a disease in a subject. The invention also provides a use of the of the polynucleotide, vector, cell, RNA molecule or pharmaceutical composition of the invention in the manufacture of a medicament for use in a method of treating or preventing a disease in a subject. The subject is typically a mammalian subject. Typically, the subject is a human. The subject may be a non-human mammal, such as a mouse, rat, cat, dog, pig, goat, sheep, horse, cow, camel or non-human primate. The disease, disorder or condition to be treated depends on the therapeutic agent encoded by the polynucleotide, vector, RNA molecule or pharmaceutical composition of the invention. The disease, disorder or condition is typically one which can be treated by the therapeutic agent encoded by the polynucleotide, vector, RNA molecule or pharmaceutical composition, for example by the antigen, peptide, polypeptide or protein encoded by the ORF, or by the encoded non-coding functional RNA. The disclosure relates to a segmented poly(A) tail which can be used in a platform and adapted to express molecules related to existing or future therapies. Accordingly, the therapeutic use is not particularly limited. For example, the disease, disorder or condition may be a viral infection, a bacterial infection, a fungal infection, a parasitic infection, cancer, an autoimmune disease, allergy, a genetic disorder or the like. The therapeutic use may be as a vaccine. mRNA vaccines are well known in the art, particularly in view of the recent SARS-CoV-2 pandemic. The therapeutic use may be as part of enzyme replacement therapy (ERT). The therapeutic use may be as part of a gene therapy. The therapeutic use may be for expression of a peptide, polypeptide or protein therapeutic molecule, for example an antibody. The quantities of peptide, polypeptide or protein for such therapeutic uses are typically much greater than that required for mRNA vaccines, and the RNA molecules of the present invention are particularly adapted to allow for robust expression of such molecules. Also described herein is a method of transforming a host cell with a polynucleotide, vector, RNA molecule or pharmaceutical composition of the invention. The polynucleotide, vector, RNA molecule or pharmaceutical composition may encode or comprise a sequence encoding a therapeutic RNA sequence, a therapeutic polynucleotide, peptide, polypeptide, protein or antigen as described herein. The host cell may be a cell for use in cellular therapy. For example, the host cell may be a stem cell, such a haematopoietic stem cell, a skeletal muscle stem cell, a mesenchymal stem cell, or a tissue-specific progenitor cell, such as a cardiac progenitor cell or a liver progenitor cell. The host cell may be a cell for use in adoptive cell therapy. For example, the host cell may be a lymphocyte, such as a T cell or a natural killer cell. The host cell may be a professional antigen presenting cell, for example a dendritic cell, a monocyte or a macrophage. The host cell may be a tissue specific cell, such as a pancreatic islet cell. Methods of propagation The invention also provides a method of propagating the polynucleotide or vector of the invention. The term “propagating” is intended to refer to the making of copies of the polynucleotide or vector, and may be used interchangeably with the term “replicating”. As discussed herein, non-segmented poly(A) tails suffer from low stability when encoded in a polynucleotide or vector. When the polynucleotide or vector is propagated, such as in an E. coli cell, the non-segmented poly(A) tail is observed to be truncated with relatively high frequency (see Figure 1). It is an aim of the invention to improve the stability of the poly(A) tail and reduce the incidence of the poly(A) tail becoming truncated following propagation. The polynucleotide or vector is typically propagated in a cell. The cell may be a prokaryotic cell, such as a bacterial cell. The bacterial cell may be an E. coli cell, such as E. coli DH5alpha. For example, the cell may be an E. coli cell comprising a mutation in the recA, recA1 and / or recA13 gene, such as deletion of the recA, recA1 and / or recA13 gene, and / or a mutation in the recBCD gene to reduce or abolish exonuclease V activity. Such cells are typically used as they efficiently propagate the polynucleotide or vector. The method may comprise transforming the cell with the polynucleotide or the vector. As used herein, the term “transforming” is intended to encompass any means by which the polynucleotide or the vector is taken up by a cell, such as by transformation, transfection or transduction. The transformation may be performed by heat shock, e.g. wherein the cell is an E.coli cell. The method comprises culturing the cell under conditions suitable for increasing the number of molecules of the polynucleotide, for example by increasing the copy number per cell and / or increasing the number of cells. The culture may be performed for 6, 12, 18, 24 or 48 hours, for example wherein the cell is an E.coli cell. The culture may be performed at 30˚C or 37˚C, for example where the cell is an E.coli cell. Where the cell is an E.coli cell, the culture may be performed by plating the E. coli (e.g. on LB agar) following transformation. The culture may be performed under conditions to select for a cell comprising the polynucleotide over cells not comprising the polynucleotide. Accordingly, the method may be a method of propagating the polynucleotide or vector of the invention in an E.coli cell, wherein the method comprises transforming the E.coli cell with the polynucleotide or the vector and culturing the transformed E.coli cell at 30˚C or 37˚C. The inventors have further identified that the stability of a poly(A) tail during propagation may be improved by propagating a polynucleotide or a vector comprising the poly(A) tail in an E.coli cell, wherein the E.coli cell is cultured at 30˚C, for example as opposed to the typical E.coli culture condition of 37˚C. According, the invention also provides a method of propagating a polynucleotide or vector in an E.coli cell, wherein the polynucleotide or vector comprises a nucleic acid sequence encoding or comprising a poly(A) tail, and wherein the method comprises transforming the E.coli cell with the polynucleotide or the vector and culturing the transformed E.coli cell at 30˚C. Any poly(A) tail may be used in this method. For example, the poly(A) tail may comprise at least 30 adenosine nucleotides, such as at least 60, at least 100, at least 120 or at least 150 adenosine nucleotides. The poly(A) tail may comprise no more than 500 adenosine nucleotides, such as no more than 300, no more than 250, or no more than 200 adenosine nucleotides. The poly(A) tail may comprise 30-300 adenosine nucleotides, such as 120- 300, 120-250, 120-200, 150-200, about 120, about 150 or about 200 adenosine nucleotides. The poly(A) tail may consist of adenosine nucleotides. Preferably, the poly(A) tail is a segmented poly(A) tail wherein the adenosine nucleotides in the poly(A) sequence are separated into segments by one or more non-adenosine nucleotides. The segmented poly(A) tail may be the segmented poly(A) tail of the invention. The segmented poly(A) tail may be a poly(A) tail described in WO 2016 / 091391 A1, WO 2020 / 074642 A1, WO 2016 / 005324 A1 or WO 2019 / 036513 A1. The method of propagation may be as further described herein, for example in respect of transformation and culture conditions. Propagation of the polynucleotide or vector of the invention may also be performed without the need for the presence of a cell. For example, the polynucleotide or vector may be propagated in the presence of a polymerase, such as a DNA-dependent DNA polymerase or an RNA-dependent RNA polymerase as appropriate, in the presence of suitable substrates (e.g. primers and nucleotide triphosphates) and under conditions to allow for the polynucleotide or vector to be propagated. For example, the polynucleotide or vector of the invention may be propagated using polymerase chain reaction (PCR) or rolling circle amplification (RCA). The method may further comprise isolating the polynucleotide or vector following propagation. The term “isolating” is intended to be used interchangeably with the term “purifying”, and refers to the separation of the polynucleotide or vector from other non- polynucleotide or vector components used in the method, such as cellular machinery, endogenous genomic polynucleotides in a cell or the like. Methods of isolating or purifying a polynucleotide or vector are well known to the skilled person. The method may further comprise determining one of more characteristics of the sequence encoding the segmented poly(A) tail in the propagated polynucleotide or vector. The purpose of this step is to evaluate the preservation of the segmented poly(A) tail following propagation. In particular, it is easier to evaluate the characteristics of the segmented poly(A) tail of the present invention when compared to segmented poly(A) tails known in the art, in part due to the segmented nature of the tail. This also allows for the length of the tail to be modified with more segments whilst retaining the ability to easily evaluate whether the correct sequence of the tail is present. The one or more characteristics may comprise the length of the segmented poly(A) tail, the number of stretches of consecutive A nucleotides in the segmented poly(A) tail, the sequence of the segmented poly(A) tail, and / or the number, identity and / or periodicity of the spacer sequences of the segmented poly(A) tail. The one or more characteristics may be determined by any means known to the skilled person, such as sequencing or gel electrophoresis. The sequencing may, for example, be Sanger sequencing, as shown in Figure 7. The step of determining one or more characteristics may comprise performing a restriction endonuclease digest of the propagated polynucleotide to thereby produce a fragment comprising the sequence encoding the segmented poly(A) tails. Suitable methods for restriction endonuclease digest of the propagated polynucleotide are provided in Examples 5 and 6. The method may further comprise analysing a fragment produced by restriction endonuclease digest of the propagated polynucleotide to determine the number of distinct stretches of consecutive A nucleotides in the segmented poly(A) tail. In vitro transcription The invention also relates to an in vitro method of producing an RNA molecule from the polynucleotide, vector, cell, or propagated polynucleotide described herein. The method may be a method of performing in vitro transcription (IVT). The method comprises contacting the polynucleotide, vector or cell of the invention, or the propagated polynucleotide obtained according to a method of the invention, with an RNA polymerase. The polynucleotide or vector is typically a DNA polynucleotide or DNA vector, and the RNA polymerase is typically a DNA-dependent RNA polymerase. The DNA-dependent RNA polymerase may be T7 RNA polymerase, SP6 RNA polymerase or T3 RNA polymerase. The polynucleotide, propagated polynucleotide or vector may be an RNA polynucleotide or RNA vector, and the RNA polymerase may be an RNA- dependent RNA polymerase. The polynucleotide, propagated polynucleotide or vector may be a circular molecule. In this case, the method typically comprises linearising the molecule prior to contact with the RNA polymerase. Linearisation of the polynucleotide, propagated polynucleotide or vector may be performed by any means known to the skilled person. Typically, the method comprises linearising the circular molecule using a restriction enzyme. Linearisation may occur at the terminal end of the segmented poly(A) tail encoded by the nucleic acid sequence. For example, the circular molecule may comprise a restriction site at the terminal end of the segmented poly(A) tail encoded by the nucleic acid sequence, and the method may comprise using a restriction enzyme to linearise the circular molecule at the restriction site. The restriction site at the terminal end of the segmented poly(A) tail encoded by the nucleic acid molecule may be a SapI restriction site, and the restriction enzyme may be SapI or an isoschizomer thereof, such as BspQI, LguI, Nt.BspQI or PciSI. The method may further comprise isolating the RNA molecule following polymerisation. As discussed above, the term “isolating” is intended to be used interchangeably with the term “purifying”, and refers to the separation of the RNA molecule from molecules or components used in the method, such as the polynucleotide or vector. For example, a DNase enzyme, such as DNaseI may be added to the reaction mixture following polymerisation to degrade the template polynucleotide or vector. The RNA molecule may then be purified using standard methods known to the skilled person, for example using ethanol precipitation-, spin column-, phenol-chloroform extraction- or bead-based extraction methods. The method may further comprise formulating the RNA molecule into a pharmaceutical composition. The pharmaceutical composition may further comprise a pharmaceutically acceptable excipient and / or a micelle, liposome, exosome or lipid nanoparticle, as described herein. The method is performed under conditions suitable for the production of an RNA molecule, for example in the presence of ribonucleotide triphosphates. The method may be performed in the presence of a 7-methyl guanosine cap nucleotide or cap analogue under conditions suitable to achieve 5’ capping of the RNA molecule. The cap may be added co-transcriptionally or post-transcriptionally. Other methods In some cases, an in cellulo or in vivo method of producing an RNA molecule from the polynucleotide, vector or propagated polynucleotide is provided. The method may comprise transfecting the polynucleotide, vector or propagated polynucleotide into a cell, and transcribing the polynucleotide, vector or propagated polynucleotide to produce the RNA molecule. The cell typically provides the conditions to allow for the production of the RNA molecule by the cell, such as an RNA polymerase. The RNA polymerase may be endogenous to the cell or exogenous to the cell. The cell is typically a eukaryotic cell. This may allow RNA modifications, such as 5’ capping, to be performed within the cell. The eukaryotic cell may be a yeast cell. The eukaryotic cell may be an animal cell, such as a mammalian cell. The mammalian cell may be a human cell or a non-human mammalian cell, such as a mouse, rat, cat, dog, pig, goat, sheep, horse, cow, camel or non-human primate cell. The cell may be an isolated cell, e.g. isolated from the mammals described above. The method typically further comprises purifying the RNA molecule from the cell. The purification may be performed by any means known to the skilled person. The purification may include a separating the RNA molecule produced by transcription of the polynucleotide, vector or propagated polynucleotide from other RNA molecules produced by the cell. In some cases, the invention also relates to a non-therapeutic and / or cosmetic use of the polynucleotide, vector or RNA molecule of the invention. For example, the invention provides a non-therapeutic and / or cosmetic method of administering the polynucleotide, vector or RNA molecule of the invention to a subject. The invention also provides a non-therapeutic and / or cosmetic use of the polynucleotide, vector or RNA molecule of the invention in a subject. The invention also provides the polynucleotide, vector or RNA molecule of the invention for a non-therapeutic and / or cosmetic use in a subject. The polynucleotide, vector or RNA typically does not comprise or encode a therapeutic molecule, such as an antigen, or a therapeutic peptide, polypeptide or protein. The polynucleotide, vector or RNA typically does not have a therapeutic use. For example, the polynucleotide, vector or RNA does not have a use for prophylaxis or treatment of a disease. In some cases, the polynucleotide, vector or RNA may have a therapeutic use, but the non-therapeutic and / or cosmetic use is restricted to use in subjects that would not have a therapeutic benefit. For example, the cosmetic use may be weight loss, and the use would be in subjects that would not benefit therapeutically from the use, for example in individuals that are not overweight. The subject is typically a mammalian subject. Typically, the subject is a human. The subject may be a non-human mammal, such as a mouse, rat, cat, dog, pig, goat, sheep, horse, cow, camel or non-human primate. The invention also provides a method of producing an RNA molecule, peptide, polypeptide, protein or antigen. The invention may be an in vitro method or an in vivo method. The method comprises transcribing or translating the RNA molecule of the invention or obtained according to the method of the invention. The RNA molecule, peptide, polypeptide, protein or antigen is typically a therapeutic RNA molecule, peptide, polypeptide, protein or antigen, as described herein. The method is typically performed in vivo following administration of the RNA molecule to a subject and incorporation of the RNA molecule into the subject’s cells. The invention also provides a method of increasing the stability or translational capacity of a polynucleotide comprising a nucleic acid sequence comprising or encoding a poly(A) tail. The method comprises replacing the sequence comprising or encoding the poly(A) tail with a sequence comprising or encoding a segmented poly(A) tail as defined herein. The method may comprise directly replacing the sequence comprising or encoding the poly(A) tail within a polynucleotide or vector. The method may comprise cloning an ORF from a vector comprising or encoding a poly(A) tail and inserting it into a polynucleotide or vector of the invention comprising a segmented poly(A) tail. Examples Example 1 – Uninterrupted adenosine homopolymers induce recombination in circular plasmids To transcribe custom-made therapeutic mRNAs, a parental plasmid vector was designed with designated 5’ and 3’ UTRs that flank the open reading frame (ORF) of the luciferase reporter gene. Downstream of the 3’UTR, sandwiched by KpnI and SapI restriction enzyme sites, the plasmid features a 120-nucleotide-long polyadenosine tract (Fig.1). The platform is flexible and each component can be exchanged using unique restriction enzymes flanking each feature. To serve as a template for in vitro transcription (IVT), the plasmid is linearised by digestion with the SapI restriction enzyme that cuts the vector at the end of the polyadenosine tract leaving a 5’ thymidine overhang. T7 RNA polymerase-transcribed mRNAs can thus be produced that feature a “clean” 120 -nucleotide-long poly(A) tail at their very 3’end (Fig.1). This parental plasmid was employed to clone new templates to produce mRNAs that encode specific therapeutic proteins. Consequently, the luciferase ORF was excised from the parental plasmid and new ORFs are subcloned to replace the luciferase ORF. Sequencing of these new subclones revealed that the plasmids contained the correct new ORFs but featured shorter adenosine homopolymer stretches ranging between 30-95 nucleotides (Fig.1). This suggested that the parental plasmid, during the subcloning process, is subject to recombination in the bacterial host, resulting in daughter plasmids featuring shorter adenosine homopolymers. The recombination-associated trimming of the adenosine homopolymers makes subcloning using such parental plasmids time- consuming and cumbersome. Furthermore, recombination presents a constant concern that during the regular plasmid amplification, required to satisfy high plasmid template demand during the scaled-up mRNA production phases, the length of the homopolymer stretches may be compromised. In addition, recombination prevents the stable production of IVT mRNAs with poly(A) tails that are longer than 120 nucleotides. This phenomenon, whilst having previously been observed (Grier et al., 2016; Trepotec et al., 2019) is not widely reported in the literature. This is surprising considering that exogenous mRNAs require poly(A) tails of at least 100 nucleotides to ensure efficient translation and longer poly(A) tails are instrumental to achieve maximum translation expression in specific tissues. Example 2 - GA27-segments facilitate cloning of extended poly(A) tracts in IVT plasmids To address this limitation, the poly(A)-encoding region in the parental plasmid was redesigned with a variant that suppresses recombination during cloning and / or amplification of plasmids. To this end, a poly(A) variant was constructed using a series of GA27segments (Fig.2). The rationale behind employing multiple GA27segments is to use the smallest possible disruptor of the adenosine homopolymer by insertion of single guanosines at specific positions. The design is further informed by the fact that CCR4- NOT appears to degrade poly(A) tails in steps of 27 adenosines (Yi et al., 2018). Single guanosines were utilised to minimise interference with PABPC1 multimerization, which is important for cap / poly(A)-dependent translation (Sawazaki et al., 2018). G residues have been shown to suppress deadenylation by CCR4-NOT activity (Lim et al., 2018) which may extend the half-life of the mRNA. It was hypothesised that the GA27-segments would stabilise the poly(A) tract in bacterial hosts, increasing the number of clones with the desired poly(A) tract length after cloning. To test this, the 114nt GA27-segmented poly(A) tail (i.e. A30GA27GA27GA27) or an uninterrupted 120nt poly(A) tract was cloned using the same parental plasmid vector containing a 30nt poly(A) tract. After transforming E. coli, 4 clones were picked at random for each poly(A) tract design and the number of each nucleotide in the poly(A) tract region was determined by sequencing (Fig.3). This showed that the GA27- segmented poly(A) tract design increased the number of clones with the desired number of adenosines: all 4 clones had close to the desired 111 adenosines. In contrast, the clones containing the uninterrupted 120nt poly(A) tract showed significantly fewer adenosines than desired, indicative of recombination. As an orthogonal technique, restriction enzyme digests of purified DNA were carried out from several clones containing either the uninterrupted 120nt poly(A) tract or the 114nt GA27-segmented poly(A) tract (Fig.4). The restriction enzymes used (BssHII and HindIII) excise a DNA fragment containing the 3’UTR and the poly(A) tract. Most of the clones expected to carry an uninterrupted 120nt poly(A) tract showed two bands in gel electrophoresis of digestion products (Fig.4A); this can be explained by recombination of the 120nt poly(A) tract, resulting in a proportion of plasmids with a shorter poly(A) tract that is visible as a shorter excised DNA fragment. In contrast, most 114nt GA27minipreps showed one defined band on the gel at the expected position (Fig.4B). This suggests that the GA27design stabilizes the poly(A) tract, suppressing recombination. As a control, BioNTech’s split poly(A) tract (A30GCATATGACTA70; Vogel et al., 2021, Stadler et al.2017) was cloned from the same original parental plasmid vector. Restriction enzyme digests of purified DNA from individual clones showed that the novel GA27-segmentation strategy performed similarly to BioNTech’s split poly(A) design (Fig.4C). After successfully producing the 114nt GA27-segmented poly(A) tract, it was tested whether the increased stability of this design would enable production of very long poly(A) tracts (>120nt). This has so far proven challenging with circular plasmids. The GA27-segmented poly(A) tract was increased from 114nt to 198nt by inserting three additional GA27segments. After transforming E. coli and picking 4 clones at random, we sequenced the poly(A) tract region, as before (Fig.3). This showed that 2 of the 4 clones contained approximately the desired number of adenosines (192 adenosines) and the other 2 clones had poly(A) tracts longer than the original 114nt. This demonstrates that GA27-segmentation is an effective strategy to produce poly(A) tracts of up to 198nt (Fig.2). Example 3 - GA27 segmentation of poly(A) tail has no impact on translation efficiency We next evaluated whether the GA27-segmented poly(A) tailed mRNAs are efficiently translated. Luciferase-encoding mRNAs with either a 114nt GA27-segmented poly(A) tail (Fig.5A) or an uninterrupted 30nt or 120nt standard poly(A) tail were transcribed in vitro. These mRNAs were transfected at equal amounts into HEK-293T cells. After 48h the cells were lysed and subjected to a luciferase activity assay. The 114nt GA27- segmented poly(A) tail had similar translational capacity as an uninterrupted 120nt poly(A) tail, demonstrating that the interruption of the adenosine homopolymers by insertions of guanosines every 27 nucleotides has no adverse impact on translation efficiency compared to mRNAs with uninterrupted poly(A) tails (Fig.5B). The translational output of the GA27-segmented poly(A) mRNAs was also compared with BioNTech’s 110nt split poly(A) tail used in the FDA-approved COVID-19 mRNA vaccine, BNT162b2 (Vogel et al., 2021). The 114nt GA27-segmented poly(A) mRNAs had similar translational capacity as equivalent mRNA featuring BioNTech’s split poly(A) tail (Fig.6). The 198-nucleotide-long poly(A) tail composed of GA27segments was also evaluated. This suggested that the extended 198-nucleotide-long poly(A) tail may further increase the translational capacity of IVT mRNA beyond that achieved with uninterrupted poly(A) tails of 120 adenosines or BioNTech’s split poly(A) tail. Example 4 - Quality control by sequencing is simplified in GA27-segmented polyadenosine plasmids An additional feature of the GA27-segmented polyadenosine plasmids is the relative ease by which the integrity of the poly(A) stretches in plasmids can be verified by sequencing. The guanosines in the GA27segments proved effective for the quality control of the plasmids as they clearly identify each segment (Fig.7) by the presence of a designated guanosine peak, allowing rapid verification of the exact length of the encoded tails. Example 5 – GA27-segmented poly(A) tract is stable during plasmid amplification and retransformation in bacterial hosts To satisfy the large DNA template demand of commercial-scale in vitro transcription of therapeutic mRNAs, plasmid DNA is typically amplified in E. coli. As the length of the poly(A) tract in the plasmid directly affects the translational output of the transcribed mRNA, it is critical that the length of the poly(A) tract does not shorten during this amplification step. To evaluate the stability of different poly(A) tract sequences in otherwise identical plasmids during amplification, the length of the poly(A) tract region was assessed in plasmids purified from a small 3 mL overnight E. coli culture (miniprep) or from a scaled-up 100 mL overnight culture inoculated from this smaller 3mL culture (midiprep). As before, the purified plasmid DNA was digested with HindIII and BssHII to excise a DNA fragment containing the 3’UTR and poly(A) tract. The fragments were subsequently separated by gel electrophoresis. For an uninterrupted 120nt poly(A) tract, the dominant band for both the miniprep and midiprep digests migrated to the expected position for a 120nt poly(A) tract. However, a fainter, more diffuse band could be seen below this band in both digests (Fig.8A). This lower diffuse band can be explained by shortening of the 120nt poly(A) tract during amplification. The presence of a shorter poly(A) tract-containing plasmid species within the miniprep and midiprep samples is also supported by the presence of non-adenosine peaks in the Sanger sequencing spectra (Fig.8A). In contrast, digestion of the miniprep or midiprep plasmids containing the GA27-segmented poly(A) tract resulted in an identical, defined band at the expected position for a 114nt poly(A) tract (Fig.8B). In addition, the Sanger sequencing spectra of both the miniprep and midiprep plasmid samples showed no non-adenosine peaks apart from the expected guanosine peaks, which appeared at the correct positions (Fig. 8B). This suggests the GA27-segmented poly(A) tract is highly stable during amplification in E. coli. As a control, the stability of BioNTech’s split poly(A) tract was evaluated and found to be similarly stable to the GA27-segmented poly(A) design (Fig. 8C). To further assess the stability of the GA27-segmented poly(A) tract in bacterial hosts, their stability during re-transformation of the plasmids in E. coli was investigated. To assess the stability of re-transformed plasmids, the plasmid DNA was purified from single transformants and digested with BssHII and HindIII as before.8 out of 10 clones maintained full-length 114nt GA27-segmented poly(A) tracts throughout the transformation and growth process as verified by gel electrophoresis (Fig.9). This further emphasises the high stability of the GA27-segmented poly(A) tract in E. coli. Example 6 – GA27 segments stabilise the poly(A) tract during subcloning It was next determined whether the GA27-segmented poly(A) tract suppresses recombination during subcloning. To that end, the luciferase ORF of the 120-adenosine- or the 114-nucleotide-long GA27-segmented poly(A) tract-containing plasmids was exchanged and 4 subclones were randomly picked each from the plated transformed bacteria. The purified plasmid DNA was digested from these 4 subclones with restriction enzymes that excise the ORF and a fragment consisting of the 3’UTR and the poly(A) tract. To determine the length of the poly(A) tract after subcloning, the digest products were separated via gel electrophoresis (Fig.10). For the uninterrupted 120nt poly(A) tract, four out of four (4 / 4) subclones had undergone clear shortening of the poly(A) tracts after subcloning as evidenced by the appearance of a strong shortened DNA fragment at around 400 bp; below the expected position of 439 bp (Fig.10, right panel). In contrast, for the GA27-segmented poly(A) tract design, only 1 / 4 subclones showed clear shortening (Fig.10, left panel). Furthermore, in the singular subclone that showed clear shortening of the poly(A) tract, sequencing revealed that one complete GA27segment was lacking. This suggests that if shortening does occur, it is due to the loss of entire GA27segments, rather than shortening within GA27segments. This characteristic of the design enables rapid identification of clones with the correct length poly(A) tract via simple restriction enzyme digests and subsequent gel electrophoresis. To compare the stability of our 114nt GA27-segmented poly(A) tract with BioNTech’s 110nt split poly(A) tract, subcloning was tested using the BioNTech poly(A) tract. This revealed that the poly(A) tract shortened in 1 / 4 subclones (Fig.11). Therefore, the GA27- segmented poly(A) tract shows similar stability to BioNTech’s poly(A) tract design during subcloning. This method was repeated to investigate the stability of the 198-nucleotide-long GA27- segmented poly(A) tract. However, this showed that 1 / 4 subclones maintained the correct length after subcloning in E. coli at 37°C and the presence of a band at 0.4kb suggested that the poly(A) tract had shortened in some proportion of the plasmids within the sample (Fig.12). Therefore, the GA27-segmented poly(A) tract design enables the construction of plasmids harboring very long poly(A) tracts but the design requires some optimization to facilitate stable subcloning of such plasmids. To further test the stability of the GA27-segmented poly(A) tract, a second ORF (ORF2) of different length and sequence composition to ORF1 was subcloned into a plasmid vector containing a 114nt GA27-segmented poly(A) tract. The purified plasmid DNA from the resultant subclones was digested with BssHII, HindIII and XhoI. Gel electrophoresis of digestion products showed that in 7 / 8 subclones that successfully inserted ORF2, the dominant band representing the DNA fragment containing the 3’UTR and poly(A) tract migrated to the correct position of a 114nt poly(A) tract (same position as the band seen in the digest of the plasmid vector used for cloning (Fig.13)). In the one subclone that showed a shortened poly(A) tract, sequencing revealed that 2 entire GA27segments were missing (Fig.13, Subclone 4). As seen with subcloning ORF1, this shows that if poly(A) shortening does occur, entire GA27segments are removed, rather than an unpredictable change in length; this enables rapid identification of subclones containing the full-length poly(A) tract. Overall, this shows that the GA27-segmented poly(A) tract design can greatly accelerate the production of customized IVT mRNAs; the laborious process of picking many clones and undergoing extensive quality control to identify a clone that maintained the correct poly(A) tract length is greatly simplified. Example 7 – GA27-segmented poly(A) tract design enables production of in vitro transcription plasmids featuring poly(A) tracts of customizable length which can exceed the length of the BioNTech split poly(A) design. In contrast to the BioNTech split poly(A) tail design, the modular nature of the GA27- segmented design enables production of a range of poly(A) tract lengths by adjusting the number of GA27segments. As the GA27-segmented design stabilised the 114nt poly(A) tract during propagation of plasmids in bacterial hosts compared to an uninterrupted poly(A) tract of similar length, we tested if this design supports the production of poly(A) tracts exceeding 114nt. Remarkably, we successfully cloned a 198nt GA27- segmented poly(A) tract into the plasmid and it was stable when the culture size was increased (from 3mL to 100mL) to produce sufficient plasmid for a midiprep (Figs.3 and 14A). This is a significant result as previous designs that attempt to stabilise the poly(A) tract in circular plasmids have not demonstrated the ability to make stable poly(A) tracts of such lengths. Furthermore, the GA27-segmented design enabled production of plasmids with poly(A) tracts of customizable length up to 198nt (Fig. 14B), emphasising the potential of the modular GA27poly(A) tract design. Example 8 - Length of 198nt GA27-segmented poly(A) tract can be maintained during subcloning. We next tested the stability of the 198nt GA27-segmented poly(A) tract during subcloning by replacing the Renilla luciferase ORF in the in vitro transcription plasmid with a larger open reading frame (ORF3). The 198nt GA27-segmented poly(A) tract (Figure 15, lane V) was successfully retained after subcloning (Figure 15, subclones 1,4,5,6 and 9) the plasmids in E. coli at 30°C. Some subclones showed poly(A) tract shortening (Figs.15, subclones 2,3,7,8 and 10) but as previously observed (Figs.9 and 13), this was by loss of entire segments. This further emphasizes how the GA27design enables rapid identification of shortening events. Example 9 - Plasmids featuring a 198nt long GA27-segmented poly(A) tract are stable during amplification in E. coli. To test the stability of the 198nt GA27-segmented poly(A) tracts during amplification in E. coli at 30°C, we compared the integrity of the 198nt GA27plasmids extracted from miniprep cultures and the corresponding midiprep cultures. We tested the amplification stability of two plasmids featuring two different ORFs (ORF3 and ORF4). For both ORFs, stable miniprep and midiprep plasmids were purified (Fig.16), confirming that the GA27poly(A) tract design supports the stable amplification of plasmids featuring a 198-nucleotide long poly(A) tract. Example 10 - In vitro-transcribed mRNA with 170nt and 198nt GA27-segmented poly(A) tails show higher translational output than equivalent mRNA with 110nt BioNTech split poly(A) tail. We tested whether increasing the length of the poly(A) tract affects the translational output of in vitro transcribed mRNAs. To that end we in vitro transcribed mRNAs encoding Renilla luciferase featuring 114 nucleotide, 170 nucleotide, or 198 nucleotide GA27-segmented poly(A) tails, an mRNA featuring the BioNTech poly(A) tail and an mRNA with a 30-nucleotide long poly(A) tail. After confirming the integrity of the transcripts (Fig.17A) the mRNAs were transfected into HEK-293T cells and luciferase activity was measured. mRNAs featuring either the 170 nucleotide or 198 nucleotide long poly(A) tail resulted in increased luciferase activity compared to mRNAs featuring shorter poly(A) tails (Fig.17B). This demonstrates that the GA27design not only enables the production of stable plasmids with long (at least up to 198 nucleotides) poly(A) tracts but that mRNAs featuring poly(A) tails longer than 110 nucleotides have enhanced translational output. Example 11 - GA27-segmented poly(A) tract design allows production of mRNAs with customizable length of poly(A) tail ranging from 58 to 198 nucleotides with each GA27 segment increasing translational output. The modular GA27-segmented poly(A) tract design enabled production of IVT plasmids featuring poly(A) tracts ranging from 58nt to 198nt (Fig.14B). To explore the relationship between each additional GA27segment and the translational capacity of IVT mRNA, we produced Renilla luciferase-encoding IVT mRNA featuring each distinct poly(A) tail length (Fig.18A) and transfected them into HEK-293T cells. This revealed a striking correlation between the number of GA27segments in the poly(A) tail of mRNAs and their respective translational output (Fig.18B). These results demonstrate that the GA27-segmented poly(A) tract design enables the production of IVT mRNAs featuring poly(A) tails of customizable lengths and further provides opportunities to tailor expression to specific needs. Example 12 - 198nt GA27-segmented poly(A) tail can enable lower mRNA dose to achieve same translational output as equivalent mRNA with a shorter 114nt poly(A) tail. We hypothesized that by using the longer poly(A) tails produced with the GA27- segmented design, lower doses of IVT mRNA may achieve the same translational output as higher doses of IVT mRNA with shorter poly(A) tail lengths. We thus transfected HEK-293T cells with Renilla luciferase-encoding mRNA featuring 114nt or 198nt GA27- segmented poly(A) tails at 3 different mRNA amounts. After 48 hours, the translational capacity of 1000ng of mRNA with a 114nt poly(A) tail was comparable to 750ng of mRNA with a 198nt poly(A) tail (Fig.19). This suggests that lower doses of IVT mRNA could be used if they feature longer poly(A) tails and this can be achieved using the GA27-segmented design. Therefore, longer poly(A) tails created using the GA27design have the potential to decrease the cost of IVT mRNA vaccines and therapies. References Alanine, D. G. W. et al., (2019). Human Antibodies that Slow Erythrocyte Invasion Potentiate Malaria-Neutralizing Antibodies. Cell, 178(1), 216-228.e21. Baer, B. W., & Kornberg, R. D. (1983). The protein responsible for the repeating structure of cytoplasmic poly(A)-ribonucleoprotein. Journal of Cell Biology, 96(3), 717– 721. Chang, H. et al., (2014). Molecular Cell Resource TAIL-seq: Genome-wide Determination of Poly(A) Tail Length and 30 End Modifications. Deo, R.C. et al., (1999). Recognition of polyadenylate RNA by the poly (A)-binding protein. Cell, 98(6), pp.835-845. Grier, A. E. et al., (2016). pEVL: A Linear Plasmid for Generating mRNA IVT Templates With Extended Encoded Poly(A) Sequences. Molecular Therapy - Nucleic Acids, 5(March), e306. Holtkamp, S. et al., (2006). Modification of antigen-encoding RNA increases stability, translational efficacy, and T-cell stimulatory capacity of dendritic cells. Blood, 108(13), 4009–4017. Hou, X et al., (2021). Lipid nanoparticles for mRNA delivery. Nat Rev Mater 6, 1078– 1094. Lim, J. et al., (2018). Mixed tailing by TENT4A and TENT4B shields mRNA from rapid deadenylation. Science, 361(6403), 701–704. Rohner, E. et al., (2022). Unlocking the promise of mRNA therapeutics. https: / / doi.org / 10.1038 / s41587-022-01491-z Sawazaki, R. et al., (2018). Characterization of the multimeric structure of poly(A)- binding protein on a poly(A) tail. Scientific Reports, 8(1), 1–13. Stadler et al., (2017). Elimination of large tumors in mice by mRNA-encoded bispecific antibodies. Nature Medicine, 23 (7), 815-817 Trepotec, Z. et al., (2019). Segmented poly(A) tails significantly reduce recombination of plasmid DNA without affecting mRNA translation efficiency or half-life. Rna, 25(4), 507–518. Vogel, A. B. et al., (2021). BNT162b vaccines protect rhesus macaques from SARS- CoV-2. Nature, 592(7853), 283–289. Yi, H. et al., (2018). PABP Cooperates with the CCR4-NOT Complex to Promote mRNA Deadenylation and Block Precocious Decay. Molecular Cell, 70(6), 1081- 1088.e5. Further embodiments 1. A polynucleotide comprising a nucleic acid sequence encoding a segmented poly(A) tail, wherein the segmented poly(A) tail comprises at least 80 A nucleotides separated by at least two spacer sequences, wherein the spacer sequences are separated by a sequence of 11-35 consecutive A nucleotides. 2. A polynucleotide comprising a nucleic acid sequence comprising a segmented poly(A) tail, wherein the polynucleotide is an RNA molecule, wherein the segmented poly(A) tail comprises at least 80 A nucleotides separated by at least two spacer sequences, wherein the spacer sequences are separated by a sequence of 11-35 consecutive A nucleotides. 3. The polynucleotide of embodiment 1 or 2, wherein the segmented poly(A) tail comprises: (a) at least around 100 A nucleotides, such as at least around 150 or at least around 200 A nucleotides; and / or (b) 80-400 A nucleotides, such as 100-300 or 100-200 A nucleotides. 4. The polynucleotide of any one of the preceding embodiments, wherein the segmented poly(A) tail comprises an initial stretch of 11-35 consecutive A nucleotides, such as an initial stretch of 22-35, 23-35, 25-35, 27-33, 29-31 or around 30 A nucleotides. 5. The polynucleotide of any one of the preceding embodiments, wherein the segmented poly(A) tail comprises at least 3 spacer sequences, such as at least 4 or at least 5 spacer sequences. 6. The polynucleotide of any one of the preceding embodiments, wherein the spacer sequences are separated by a sequence of 22-35 consecutive A nucleotides, such as a sequence of 22-30, 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides, and / or wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 or 35 nucleotides. 7. A polynucleotide comprising a nucleic acid sequence encoding a segmented poly(A) tail, wherein the segmented poly(A) tail comprises a sequence of the formula AX(SAY)z, wherein S is a spacer sequence, X is 11-35, Y is 11-35 and Z is at least 2. 8. A polynucleotide comprising a nucleic acid sequence comprising a segmented poly(A) tail, wherein the polynucleotide is an RNA molecule, wherein the segmented poly(A) tail comprises a sequence of the formula AX(SAY)z, wherein S is a spacer sequence, X is 11-35, Y is 11-35 and Z is at least 2. 9. The polynucleotide of embodiment 7 or 8, wherein X is 22-35, such as 23-35, 25- 35, 27-33, 29-31 or around 30. 10. The polynucleotide of any one of embodiments 7 to 9, wherein Y is 22-35, such as 22-30, 23-30, 25-29, 26-28 or around 27. 11. The polynucleotide of any one of embodiments 7 to 10, wherein Z is at least 3, such as at least 4 or at least 5. 12. The polynucleotide of any one of embodiments 7 to 11, wherein the segmented poly(A) tail comprises a sequence of the formula AX(SAY)zAN, wherein N is: (a) at least 10; (b) at most 80; and / or (c) 22-35, 44-70, 66-105 or 88-120. 13. The polynucleotide of any one of the preceding embodiments, wherein each spacer sequence is 1-25 nucleotides in length, such as 1-12 or 1-6 nucleotides in length. 14. The polynucleotide of any one of the preceding embodiments, wherein each spacer sequence is a single nucleotide. 15. The polynucleotide of any one of the preceding embodiments, wherein each spacer sequence does not comprise an A nucleotide at the first or last position of the spacer. 16. The polynucleotide of any one of the preceding embodiments, wherein each spacer sequence does not comprise an A nucleotide. 17. The polynucleotide of any one of the preceding embodiments, wherein each spacer sequence consists of B nucleotides (i.e. C, G or T). 18. The polynucleotide of any one of the preceding embodiments, wherein each spacer sequence consists of a single B nucleotide (i.e. C, G or T). 19. The polynucleotide of any one of the preceding embodiments, wherein each spacer sequence consists of K nucleotides (i.e. G or T). 20. The polynucleotide of any one of the preceding embodiments, wherein each spacer sequence consists of a single K nucleotide (i.e. G or T). 21. The polynucleotide of any one of the preceding embodiments, wherein each spacer sequence consists of G nucleotides. 22. The polynucleotide of any one of the preceding embodiments, wherein each spacer sequence consists of a single G nucleotide. 23. The polynucleotide of any one of the preceding embodiments, wherein the segmented poly(A) tail is at the 3’ end of the sequence encoded by the nucleic acid sequence or the 3’ end of the nucleic acid sequence. 24. The polynucleotide of any one of the preceding embodiments, wherein the sequence encoded by the nucleic acid sequence or the nucleic acid sequence further comprises: (i) a 5’ untranslated region (5’ UTR); (ii) an open reading frame (ORF), a sequence for introducing an ORF, or a sequence encoding a peptide, polypeptide or protein; and / or (iii) a 3’ untranslated region (3’ UTR). 25. The polynucleotide of any one of the preceding embodiments, wherein the sequence encoded by the nucleic acid sequence or the nucleic acid sequence comprises in order: (i) a 5’ untranslated region (5’ UTR); (ii) an open reading frame (ORF), a sequence for introducing an ORF, or a sequence encoding a peptide, polypeptide or protein; (iii) a 3’ untranslated region (3’ UTR); and (iv) the segmented poly(A) tail. 26. The polynucleotide of embodiment 24 or 25, wherein the ORF encodes an antigen and / or a therapeutic peptide, polypeptide or protein. 27. The polynucleotide of any one of the preceding embodiments, wherein the sequence encoded by the nucleic acid sequence or the nucleic acid sequence further comprises a non-coding functional RNA sequence. 28. The polynucleotide of any one of the preceding embodiments, wherein the sequence encoded by the nucleic acid sequence or the nucleic acid sequence comprises in order: (i) a non-coding functional RNA sequence; and (ii) the segmented polyA tail. 29. The polynucleotide of any one of the preceding embodiments, wherein the sequence encoded by the nucleic acid sequence or the nucleic acid sequence further comprises an internal ribosome entry site (IRES). 30. The polynucleotide of any one of embodiments 1, 3 to 7 and 9 to 29, wherein the polynucleotide further comprises a promoter, such as the T7, SP6 or T3 promoter sequence. 31. The polynucleotide of any one of embodiments 1, 3 to 7 and 9 to 30, wherein the polynucleotide further comprises: (i) a marker gene; (ii) an origin of replication; (iii) a multiple cloning site; and / or (iv) a restriction site, preferably at the terminal end of the segmented poly(A) tail encoded by the nucleic acid sequence, optionally wherein the restriction site is a SapI restriction site. 32. The polynucleotide of any one of embodiments 1, 3 to 7 and 9 to 31, which is a DNA polynucleotide. 33. The polynucleotide of any one of embodiments 1, 3 to 7 and 9 to 32, wherein the sequence encoding the segmented poly(A) tail exhibits higher stability than a control polynucleotide encoding a non-segmented poly(A) tail comprising the same number of A nucleotides, wherein stability is calculated following propagation of the polynucleotide in Escherichia coli. 34. The polynucleotide of any one of the preceding embodiments, wherein the nucleic acid sequence encodes or corresponds to an RNA molecule having the segmented poly(A) tail, wherein the RNA molecule has the same or greater translational capacity than a control RNA molecule comprising a non-segmented poly(A) tail comprising the same number of A nucleotides, wherein translational capacity is calculated as the measured level of luciferase expressed 48 hours after transformation of the RNA molecule or control RNA molecule into a HEK293T host cell, where the RNA molecule and the control RNA molecule each comprise a sequence encoding a luciferase. 35. A vector comprising the polynucleotide of any one of the preceding embodiments. 36. The vector of embodiment 35, which is a linear vector or a circular vector. 37. A cell comprising the polynucleotide of any one of embodiments 1 to 34, or the vector of embodiment 35 or 36. 38. The cell of embodiment 37, which is a prokaryotic cell, preferably a bacterial cell, more preferably an Escherichia coli cell. 39. An RNA molecule obtained or obtainable by transcription of, or encoded by, the nucleic acid sequence of any one of embodiments 1 to 34, or the vector of embodiment 35 or 36. 40. The polynucleotide or RNA molecule of any one of embodiments 2 to 6, 8 to 29, 34 and 39, which is an mRNA molecule. 41. The polynucleotide or RNA molecule of any one of embodiments 2 to 6, 8 to 29, 34, 39 and 40, which comprises a 5’ cap, preferably wherein the 5’ cap comprises a 7- methylguanosine cap. 42. A pharmaceutical composition comprising the polynucleotide or RNA molecule of any one of embodiments 2 to 6, 8 to 29, 34, and 39 to 41. 43. The pharmaceutical composition of embodiment 42, further comprising a pharmaceutically acceptable excipient. 44. The pharmaceutical composition of embodiment 42 or 43, further comprising a micelle, liposome, exosome or lipid nanoparticle. 45. A method of treating or preventing a disease, disorder or condition in a subject, the method comprising administering the pharmaceutical composition of any one of embodiments 42 to 44 to the subject. 46. The pharmaceutical composition of any one of embodiments 42 to 44 for use in a method of treating or preventing a disease in a subject. 47. Use of the pharmaceutical composition of any one of embodiments 42 to 44 in a method of treating or preventing a disease in a subject. 48. Use of the polynucleotide of any one of embodiments 1 to 34, the vector of embodiment 35 or 36, the cell of embodiment 37 or 38, the polynucleotide or RNA molecule of any one of embodiments 39 to 41, or the pharmaceutical composition of any one of embodiments 42 to 44 in the manufacture of a medicament for use in a method of treating or preventing a disease in a subject. 49. A method of propagating a polynucleotide, comprising propagating the polynucleotide of any one of embodiments 1 to 34, or the vector of embodiment 35 or 36, in a cell. 50. The method of embodiment 49, wherein the cell is a prokaryotic cell, preferably a bacterial cell, more preferably an Escherichia coli cell. 51. The method of embodiment 49 or 50, which comprises transforming the cell with the polynucleotide of any one of embodiments 1 to 34 or the vector of embodiment 35 or 36. 52. The method of any one of embodiments 49 to 51, which further comprises isolating the polynucleotide or the vector following propagation. 53. The method of any one of embodiments 49 to 52, which further comprises determining one or more characteristics of the sequence encoding the segmented poly(A) tail in the propagated polynucleotide or vector. 54. The method of embodiment 53, wherein the one or more characteristics comprise (i) the length of the segmented poly(A) tail, (ii) the number of stretches of consecutive A nucleotides in the segmented poly(A) tail, and / or (iii) the sequence of the segmented poly(A) tail. 55. The method of embodiment 53 or 54, wherein the one or more characteristics are determined by sequencing and / or gel electrophoresis. 56. The method of any one of embodiments 53 to 55, which comprises performing a restriction digest of the propagated polynucleotide to thereby produce a fragment comprising the sequence encoding the segmented poly(A) tail. 57. The method of embodiment 56, further comprising analysing the fragment by electrophoresis to determine the number of distinct stretches of consecutive A nucleotides in the segmented poly(A) tail. 58. An in vitro method of producing an RNA molecule, comprising contacting the polynucleotide of any one of embodiments 1 to 34, the vector of any one of embodiments 35 or 36, the cell of embodiment 37 or 38, or the propagated polynucleotide obtained by the method of any one of embodiments 49 to 57, with an RNA polymerase. 59. The method of embodiment 58, wherein the polynucleotide, propagated polynucleotide or vector is a circular molecule, and the method further comprises linearising the circular molecule. 60. The method of embodiment 59, wherein linearising the circular molecule is carried out using a restriction enzyme. 61. The method of embodiment 59 or 60, wherein linearising the circular molecule occurs at the terminal end of the segmented poly(A) tail encoded by the nucleic acid sequence. 62. The method of embodiment 61, wherein the polynucleotide, propagated polynucleotide or vector comprises a SapI restriction site at the terminal end of the segmented poly(A) tail encoded by the nucleic acid sequence and the restriction enzyme is SapI or an isoschizomer thereof. 63. The method of any one of embodiments 58 to 62, wherein the RNA polymerase is a DNA-dependent RNA polymerase. 64. The method of any one of embodiments 58 to 63, wherein the RNA polymerase is T7 RNA polymerase, SP6 RNA polymerase or T3 RNA polymerase. 65. The method of any one of embodiments 58 to 64, which further comprises isolating the RNA molecule following the step of contacting the polynucleotide, vector or cell with the RNA polymerase. 66. The method of any one of embodiments 58 to 65, which further comprises formulating the RNA molecule into a pharmaceutical composition. 67. The method of any one of embodiments 58 to 66, wherein the pharmaceutical composition further comprises a pharmaceutically acceptable excipient, and / or a micelle, liposome, exosome or lipid nanoparticle. 68. A pharmaceutical composition produced by the method of embodiment 66 or 67. 69. An in vitro or in vivo method of producing a peptide, polypeptide or protein, the method comprising translating the polynucleotide or RNA molecule of any one of embodiments 2 to 6, 8 to 29, 34, and 39 to 41, or obtained by the method of any one of embodiments 58 to 67. 70. A method of increasing the stability or translational capacity of a polynucleotide comprising a nucleic acid sequence encoding or comprising a poly(A) tail, comprising replacing the sequence encoding or comprising the poly(A) tail with a sequence encoding or comprising a segmented poly(A) tail as defined in any one of embodiments 1 to 34. 71. The polynucleotide of any one of embodiments 1 to 34, wherein the segmented poly(A) tail comprises at least 120 A nucleotides. 72. The polynucleotide of any one of embodiments 1 to 34 and 71, wherein the segmented poly(A) tail comprises up to 400 A nucleotides, such as up to 300 or up to 200 A nucleotides. 73. The polynucleotide of any one of embodiments 7 to 34 and 71 to 72, wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides. 74. The polynucleotide of embodiments 73, wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 35 nucleotides. 75. The polynucleotide of any one of embodiments 1 to 34 and 71 to 74, wherein the nucleic acid sequence encodes or corresponds to an RNA molecule having the segmented poly(A) tail, wherein the RNA molecule has the same or greater translational capacity than a control RNA molecule comprising a non-segmented poly(A) tail comprising the same number of A nucleotides, wherein translational capacity is calculated as the measured level of luciferase expressed 48 hours after transformation into a HEK293T host cell of a nucleic acid sequence comprising (i) the RNA molecule or the control RNA molecule and (ii) a sequence encoding a luciferase.

Claims

CLAIMS 1. A polynucleotide comprising a nucleic acid sequence encoding a segmented poly(A) tail, wherein the segmented poly(A) tail comprises at least 120 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22-35 consecutive A nucleotides, and wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides.

2. A polynucleotide comprising a nucleic acid sequence comprising a segmented poly(A) tail, wherein the polynucleotide is an RNA molecule, wherein the segmented poly(A) tail comprises at least 120 A nucleotides separated by at least three spacer sequences, wherein the spacer sequences are separated by a sequence of 22-35 consecutive A nucleotides, and wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides.

3. The polynucleotide of claim 1 or 2, wherein: (a) the segmented poly(A) tail comprises at least around 150 or at least around 200 A nucleotides; (b) the segmented poly(A) tail comprises up to 400 A nucleotides, such as up to 300 or up to 200 A nucleotides; (c) the segmented poly(A) tail comprises an initial stretch of 22-35, 23-35, 25-35, 27-33, 29-31 or around 30 consecutive A nucleotides; (d) the segmented poly(A) tail comprises at least 4 or at least 5 spacer sequences; (e) the spacer sequences are separated by a sequence of 22-30 consecutive A nucleotides, such as a sequence of 23-30, 25-29, 26-28 or around 27 consecutive A nucleotides; and / or (f) the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 35 nucleotides.

4. A polynucleotide comprising a nucleic acid sequence encoding a segmented poly(A) tail, wherein the segmented poly(A) tail comprises a sequence of the formula AX(SAY)z, wherein S is a spacer sequence, X is 22-35, Y is 22-35 and Z is at least 4, and wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides.

5. A polynucleotide comprising a nucleic acid sequence comprising a segmented poly(A) tail, wherein the polynucleotide is an RNA molecule, wherein the segmented poly(A) tail comprises a sequence of the formula AX(SAY)z, wherein S is a spacer sequence, X is 22-35, Y is 22-35 and Z is at least 4, and wherein the segmented poly(A) tail comprises no stretch of consecutive A nucleotides longer than 40 nucleotides.

6. The polynucleotide of claim 4 or 5, wherein: (a) X is 23-35, such as 25-35, 27-33, 29-31 or around 30; (b) Y is 22-30, such as 23-30, 25-29, 26-28 or around 27; (c) Z is at least 5; and / or (d) each spacer sequence is 1-25 nucleotides in length, such as 1-12 or 1-6 nucleotides in length.

7. The polynucleotide of any one of the preceding claims, wherein each spacer sequence: (a) is a single nucleotide; (b) does not comprise an A nucleotide at the first or last position of the spacer; (c) does not comprise an A nucleotide; (d) consists of B nucleotides (i.e. C, G or T); (e) consists of a single B nucleotide (i.e. C, G or T); (f) consists of K nucleotides (i.e. G or T); (g) consists of a single K nucleotide (i.e. G or T);(h) consists of G nucleotides; and / or (i) consists of a single G nucleotide.

8. The polynucleotide of any one of the preceding claims, wherein the segmented poly(A) tail is at the 3’ end of the sequence encoded by the nucleic acid sequence or the 3’ end of the nucleic acid sequence.

9. The polynucleotide of any one of the preceding claims, wherein the sequence encoded by the nucleic acid sequence or the nucleic acid sequence: (a) further comprises (i) a 5’ untranslated region (5’ UTR); (ii) an open reading frame (ORF), a sequence for introducing an ORF, or a sequence encoding a peptide, polypeptide or protein, optionally wherein the ORF encodes an antigen and / or a therapeutic peptide, polypeptide or protein; and / or (iii) a 3’ untranslated region (3’ UTR); (b) comprises in order: (i) a 5’ untranslated region (5’ UTR); (ii) an open reading frame (ORF), a sequence for introducing an ORF, or a sequence encoding a peptide, polypeptide or protein, optionally wherein the ORF encodes an antigen and / or a therapeutic peptide, polypeptide or protein; (iii) a 3’ untranslated region (3’ UTR); and (iv) the segmented poly(A) tail; (c) further comprises a non-coding functional RNA sequence; (d) comprises in order: (i) a non-coding functional RNA sequence; and (ii) the segmented polyA tail; and / or (e) further comprises an internal ribosome entry site (IRES).

10. The polynucleotide of any one of claims 1, 3-4 and 6-9, wherein: (a) the polynucleotide further comprises (i) a promoter, such as the T7, SP6 or T3 promoter sequence, (ii) a marker gene, (iii) an origin of replication, (iv) a multiplecloning site, and / or (v) a restriction site, preferably at the terminal end of the segmented poly(A) tail encoded by the nucleic acid sequence, optionally wherein the restriction site is a SapI restriction site; (b) the polynucleotide is a DNA polynucleotide; and / or (c) the sequence encoding the segmented poly(A) tail exhibits higher stability than a control polynucleotide encoding a non-segmented poly(A) tail comprising the same number of A nucleotides, wherein stability is calculated following propagation of the polynucleotide in Escherichia coli.

11. The polynucleotide of any one of the preceding claims, wherein the nucleic acid sequence encodes or corresponds to an RNA molecule having the segmented poly(A) tail, wherein the RNA molecule has the same or greater translational capacity than a control RNA molecule comprising a non-segmented poly(A) tail comprising the same number of A nucleotides, wherein translational capacity is calculated as the measured level of luciferase expressed 48 hours after transformation into a HEK293T host cell of a nucleic acid sequence comprising (i) the RNA molecule or control RNA molecule and (ii) a sequence encoding a luciferase.

12. A vector comprising the polynucleotide of any one of the preceding claims, optionally which is a linear vector or a circular vector.

13. A cell comprising the polynucleotide of any one of claims 1 to 11, or the vector of claim 12, optionally which is a prokaryotic cell, preferably a bacterial cell, more preferably an Escherichia coli cell.

14. An RNA molecule obtained or obtainable by transcription of, or encoded by, the nucleic acid sequence of any one of claims 1 to 11, or the vector of claim 12.

15. The polynucleotide or RNA molecule of any one of claims 2-3, 5-9, 11 and 14, which: (a) is an mRNA molecule; and / or(b) comprises a 5’ cap, preferably wherein the 5’ cap comprises a 7- methylguanosine cap.

16. A pharmaceutical composition comprising the polynucleotide or RNA molecule of any one of claims 2-3, 5-9, 11, 14 and 15.

17. The pharmaceutical composition of claim 16, further comprising: (a) a pharmaceutically acceptable excipient; and / or (b) a micelle, liposome, exosome or lipid nanoparticle.

18. The pharmaceutical composition of claim 16 or 17 for use in a method of treating or preventing a disease in a subject.

19. A method of propagating a polynucleotide, comprising propagating the polynucleotide of any one of claims 1 to 11, or the vector of claim 12, in a cell.

20. The method of claim 19, wherein: (a) the cell is a prokaryotic cell, preferably a bacterial cell, more preferably an Escherichia coli cell; (b) the method comprises transforming the cell with the polynucleotide of any one of claims 1 to 11 or the vector of claim 12; (c) the method further comprises isolating the polynucleotide or the vector following propagation; and / or (d) the method further comprises determining one or more characteristics of the sequence encoding the segmented poly(A) tail in the propagated polynucleotide or vector, optionally wherein: (i) the one or more characteristics comprise (A) the length of the segmented poly(A) tail, (B) the number of stretches of consecutive A nucleotides in the segmented poly(A) tail, and / or (C) the sequence of the segmented poly(A) tail; (ii) the one or more characteristics are determined by sequencing and / or gel electrophoresis; and / or(iii) the method comprises performing a restriction digest of the propagated polynucleotide to thereby produce a fragment comprising the sequence encoding the segmented poly(A) tail, optionally wherein the method further comprises analysing the fragment by electrophoresis to determine the number of distinct stretches of consecutive A nucleotides in the segmented poly(A) tail.

21. An in vitro method of producing an RNA molecule, comprising contacting the polynucleotide of any one of claims 1 to 11, the vector of claim 12, the cell of claim 13, or the propagated polynucleotide obtained by the method of claim 19 or 20, with an RNA polymerase.

22. The method of claim 21, wherein: (a) the polynucleotide, propagated polynucleotide or vector is a circular molecule, and the method further comprises linearising the circular molecule, optionally wherein linearising the circular molecule: (i) is carried out using a restriction enzyme; and / or (ii) occurs at the terminal end of the segmented poly(A) tail encoded by the nucleic acid sequence, optionally wherein the polynucleotide, propagated polynucleotide or vector comprises a SapI restriction site at the terminal end of the segmented poly(A) tail encoded by the nucleic acid sequence and the restriction enzyme is SapI or an isoschizomer thereof; (b) the RNA polymerase is a DNA-dependent RNA polymerase; (c) the RNA polymerase is T7 RNA polymerase, SP6 RNA polymerase or T3 RNA polymerase; (d) the method further comprises isolating the RNA molecule following the step of contacting the polynucleotide, vector or cell with the RNA polymerase; (e) the method further comprises formulating the RNA molecule into a pharmaceutical composition; and / or (f) the pharmaceutical composition further comprises a pharmaceutically acceptable excipient, and / or a micelle, liposome, exosome or lipid nanoparticle.

23. A pharmaceutical composition produced by the method of claim 22.

24. An in vitro or in vivo method of producing a peptide, polypeptide or protein, the method comprising translating the polynucleotide or RNA molecule of any one of claims 2-3, 5-9, 11, 14 and 15, or obtained by the method of claim 21 or 22.

25. A method of increasing the stability or translational capacity of a polynucleotide comprising a nucleic acid sequence encoding or comprising a poly(A) tail, comprising replacing the sequence encoding or comprising the poly(A) tail with a sequence encoding or comprising a segmented poly(A) tail as defined in any one of claims 1 to 11.

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